http://www.mdpi.com/journal/viruses Latest open access articles published in Viruses at http://www.mdpi.com/journal/viruses http://www.mdpi.com/1999-4915/5/5/1346 Kaposi’s sarcoma-associated herpesvirus is an oncogenic γ-herpesvirus that causes latent infection in humans. In cells, the viral genome adopts a highly organized chromatin structure, which is controlled by a wide variety of cellular and viral chromatin regulatory factors. In the past few years, interrogation of the chromatinized KSHV genome by whole genome-analyzing tools revealed that the complex chromatin landscape spanning the viral genome in infected cells has important regulatory roles during the viral life cycle. This review summarizes the most recent findings regarding the role of histone modifications, histone modifying enzymes, DNA methylation, microRNAs, non-coding RNAs and the nuclear organization of the KSHV epigenome in the regulation of latent and lytic viral gene expression programs as well as their connection to KSHV-associated pathogenesis. Viruses 2013-05-23 5 5 Review 10.3390/v5051346 1346 1373 1999-4915 2013-05-23 doi: 10.3390/v5051346 Zsolt Toth Kevin Brulois Jae Jung http://www.mdpi.com/1999-4915/5/5/1325 Cytomegalovirus (CMV) gene expression is repressed in latency due to heterochromatinization of viral genomes. In murine CMV (MCMV) latently infected mice, viral genomes are bound to histones with heterochromatic modifications, to enzymes that mediate these modifications, and to adaptor proteins that may recruit co-repressor complexes. Kinetic analyses of repressor binding show that these repressors are recruited at the earliest time of infection, suggesting that latency may be the default state. Kidney transplantation leads to epigenetic reprogramming of latent viral chromatin and reactivation of immediate early gene expression. Inflammatory signaling pathways, which activate transcription factors that regulate the major immediate early promoter (MIEP), likely mediate the switch in viral chromatin. Viruses 2013-05-23 5 5 Article 10.3390/v5051325 1325 1345 1999-4915 2013-05-23 doi: 10.3390/v5051325 Xue-feng Liu Xueqiong Wang Shixian Yan Zheng Zhang Michael Abecassis Mary Hummel http://www.mdpi.com/1999-4915/5/5/1292 Hepatitis C virus (HCV) is a leading cause of chronic liver disease, including chronic hepatitis, fibrosis, cirrhosis, and hepatocellular carcinoma. Hepatitis C infection associates with lipid and lipoprotein metabolism disorders such as hepatic steatosis, hypobetalipoproteinemia, and hypocholesterolemia. Furthermore, virus production is dependent on hepatic very-low-density lipoprotein (VLDL) assembly, and circulating virions are physically associated with lipoproteins in complexes termed lipoviral particles. Evidence has indicated several functional roles for the formation of these complexes, including co-opting of lipoprotein receptors for attachment and entry, concealing epitopes to facilitate immune escape, and hijacking host factors for HCV maturation and secretion. Here, we review the evidence surrounding pathogenesis of the hepatitis C infection regarding lipoprotein engagement, cholesterol and triglyceride regulation, and the molecular mechanisms underlying these effects. Viruses 2013-05-23 5 5 Review 10.3390/v5051292 1292 1324 1999-4915 2013-05-23 doi: 10.3390/v5051292 Daniel Felmlee Mohamed Hafirassou Mathieu Lefevre Thomas Baumert Catherine Schuster http://www.mdpi.com/1999-4915/5/5/1272 Successful infection of herpes simplex virus is dependent upon chromatin modulation by the cellular coactivator host cell factor-1 (HCF-1). This review focuses on the multiple chromatin modulation components associated with HCF-1 and the chromatin-related dynamics mediated by this coactivator that lead to the initiation of herpes simplex virus (HSV) immediate early gene expression. Viruses 2013-05-22 5 5 Review 10.3390/v5051272 1272 1291 1999-4915 2013-05-22 doi: 10.3390/v5051272 Jodi Vogel Thomas Kristie http://www.mdpi.com/1999-4915/5/5/1261 The role of hepatitis B virus (HBV) X protein (HBx) in the regulation of HBV replication remains controversial. In the present study, the role of HBx in regulating HBV replication was initially investigated in both HepG2 and Huh7 in vitro cell lines with a transient transfection system. Next, the regions of HBx responsible for transcriptional transactivation and promotion of HBV replication were mapped in an HBV replication mouse model by in vivo transfection of a series of HBx expression plasmids. In an in vitro setting, HBx deficiency had little effect on HBV replication in Huh7 cells, but impaired HBV replication in HepG2 cells. In an in vivo setting, HBx had a strong enhancing effect on HBV transcription and replication. For the C-terminal two-thirds of the protein (amino acids [aa] 51 to 154) was required for this function of HBx, and the regions spanning aa 52 to 72 and 88 to 154 were found to be important for the stimulatory function of HBx on HBV replication. In conclusion, the role of HBx in HBV replication regulation is affected by host cell type, and HBx has an important role in stimulating HBV transcription and replication in hepatocytes in vivo. Further, the transcriptional transactivation function of HBx may be crucial for its stimulatory effect on HBV transcription and replication. Viruses 2013-05-22 5 5 Article 10.3390/v5051261 1261 1271 1999-4915 2013-05-22 doi: 10.3390/v5051261 Dao-Yin Gong En-Qiang Chen Fei-Jun Huang Xiao-Hua Leng Xing Cheng Hong Tang http://www.mdpi.com/1999-4915/5/5/1250 Coronaviruses infect a variety of mammalian and avian species and cause serious diseases in humans, cats, mice, and birds in the form of severe acute respiratory syndrome (SARS), feline infectious peritonitis (FIP), mouse hepatitis, and avian infectious bronchitis, respectively. No effective vaccine or treatment has been developed for SARS-coronavirus or FIP virus, both of which cause lethal diseases. It has been reported that a cyclophilin inhibitor, cyclosporin A (CsA), could inhibit the replication of coronaviruses. CsA is a well-known immunosuppressive drug that binds to cellular cyclophilins to inhibit calcineurin, a calcium-calmodulin-activated serine/threonine-specific phosphatase. The inhibition of calcineurin blocks the translocation of nuclear factor of activated T cells from the cytosol into the nucleus, thus preventing the transcription of genes encoding cytokines such as interleukin-2. Cyclophilins are peptidyl-prolyl isomerases with physiological functions that have been described for many years to include chaperone and foldase activities. Also, many viruses require cyclophilins for replication; these include human immunodeficiency virus, vesicular stomatitis virus, and hepatitis C virus. However, the molecular mechanisms leading to the suppression of viral replication differ for different viruses. This review describes the suppressive effects of CsA on coronavirus replication. Viruses 2013-05-22 5 5 Review 10.3390/v5051250 1250 1260 1999-4915 2013-05-22 doi: 10.3390/v5051250 Yoshikazu Tanaka Yuka Sato Takashi Sasaki http://www.mdpi.com/1999-4915/5/5/1231 The role of enzymes involved in polycomb repression of gene transcription has been studied extensively in human cancer. Polycomb repressive complexes mediate oncogene-induced senescence, a principal innate cell-intrinsic tumor suppressor pathway that thwarts expansion of cells that have suffered oncogenic hits. Infections with human cancer viruses including human papillomaviruses (HPVs) and Epstein-Barr virus can trigger oncogene-induced senescence, and the viruses have evolved strategies to abrogate this response in order to establish an infection and reprogram their host cells to establish a long-term persistent infection. As a consequence of inhibiting polycomb repression and evading oncogene induced-senescence, HPV infected cells have an altered epigenetic program as evidenced by aberrant homeobox gene expression. Similar alterations are frequently observed in non-virus associated human cancers and may be harnessed for diagnosis and therapy. Viruses 2013-05-14 5 5 Review 10.3390/v5051231 1231 1249 1999-4915 2013-05-14 doi: 10.3390/v5051231 Margaret McLaughlin-Drubin Karl Munger http://www.mdpi.com/1999-4915/5/5/1219 The Hepatitis C virus causes chronic infections in humans, which can develop to liver cirrhosis and hepatocellular carcinoma. The Bovine viral diarrhea virus is used as a surrogate model for antiviral assays for the HCV. From marine invertebrates and microorganisms isolated from them, extracts were prepared for assessment of their possible antiviral activity. Of the 128 tested, 2 were considered active and 1 was considered promising. The best result was obtained from the extracts produced from the Bacillus sp. isolated from the sponge Petromica citrina. The extracts 555 (500 µg/mL, SI>18) and 584 (150 µg/mL, SI 27) showed a percentage of protection of 98% against BVDV, and the extract 616, 90% of protection. All of them showed activity during the viral adsorption. Thus, various substances are active on these studied organisms and may lead to the development of drugs which ensure an alternative therapy for the treatment of hepatitis C. Viruses 2013-04-29 5 5 Article 10.3390/v5051219 1219 1230 1999-4915 2013-04-29 doi: 10.3390/v5051219 Juliana Bastos Luciana Kohn Fabiana Fantinatti-Garboggini Marina Padilla Eduardo Flores Bárbara da Silva Cláudia de Menezes Clarice Arns http://www.mdpi.com/1999-4915/5/5/1208 REST is a key component of the HDAC1 or 2, CoREST, LSD1, REST (HCLR) repressor complex. The primary function of the HCLR complex is to silence neuronal genes in non-neuronal cells. HCLR plays a role in regulating the expression of viral genes in productive infections as a donor of LDS1 for expression of α genes and as a repressor of genes expressed later in infection. In sensory neurons the HCLR complex is involved in the silencing of viral genome in the course of establishment of latency. The thesis of this article is that (a) sensory neurons evolved a mechanism to respond to the presence and suppress the transmission of infectious agents from the periphery to the CNS and (b) HSV evolved subservience to the HCLR with at least two objectives: to maintain a level of replication consistent with maximal person-to-person spread and to enable it to take advantage of neuronal innate immune responses to survive and be available for reactivation shielded from adaptive immune responses of the host. Viruses 2013-04-29 5 5 Review 10.3390/v5051208 1208 1218 1999-4915 2013-04-29 doi: 10.3390/v5051208 Guoying Zhou Te Du Bernard Roizman http://www.mdpi.com/1999-4915/5/4/1175 Small ruminant lentiviruses (SRLV) cause a multisystemic chronic disease affecting animal production and welfare. SRLV infections are spread across the world with the exception of Iceland. Success in controlling SRLV spread depends largely on the use of appropriate diagnostic tools, but the existence of a high genetic/antigenic variability among these viruses, the fluctuant levels of antibody against them and the low viral loads found in infected individuals hamper the diagnostic efficacy. SRLV have a marked in vivo tropism towards the monocyte/macrophage lineage and attempts have been made to identify the genome regions involved in tropism, with two main candidates, the LTR and env gene, since LTR contains primer binding sites for viral replication and the env-encoded protein (SU ENV), which mediates the binding of the virus to the host’s cell and has hypervariable regions to escape the humoral immune response. Once inside the host cell, innate immunity may interfere with SRLV replication, but the virus develops counteraction mechanisms to escape, multiply and survive, creating a quasi-species and undergoing compartmentalization events. So far, the mechanisms of organ tropism involved in the development of different disease forms (neurological, arthritic, pulmonary and mammary) are unknown, but different alternatives are proposed. This is an overview of the current state of knowledge on SRLV genetic variability and its implications in tropism as well as in the development of alternative diagnostic assays. Viruses 2013-04-23 5 4 Review 10.3390/v5041175 1175 1207 1999-4915 2013-04-23 doi: 10.3390/v5041175 Hugo Ramírez Ramsés Reina Beatriz Amorena Damián Andrés Humberto Martínez http://www.mdpi.com/1999-4915/5/4/1153 Starting in late November 2010, the H5N1 highly pathogenic avian influenza (HPAI) virus was isolated from many types of wild ducks and raptors and was subsequently isolated from poultry in Korea. We assessed the genetic and pathogenic properties of the HPAI viruses isolated from a fecal sample from a mandarin duck and a dead Eurasian eagle owl, the most affected wild bird species during the 2010/2011 HPAI outbreak in Korea. These viruses have similar genetic backgrounds and exhibited the highest genetic similarity with recent Eurasian clade 2.3.2.1 HPAI viruses. In animal inoculation experiments, regardless of their originating hosts, the two Korean isolates produced highly pathogenic characteristics in chickens, ducks and mice without pre-adaptation. These results raise concerns about veterinary and public health. Surveillance of wild birds could provide a good early warning signal for possible HPAI infection in poultry as well as in humans. Viruses 2013-04-23 5 4 Article 10.3390/v5041153 1153 1174 1999-4915 2013-04-23 doi: 10.3390/v5041153 Jun-Gu Choi Hyun-Mi Kang Woo-Jin Jeon Kang-Seuk Choi Kwang-Il Kim Byung Song Hee-Soo Lee Jae-Hong Kim Youn-Jeong Lee http://www.mdpi.com/1999-4915/5/4/1143 Human Immunodeficiency Virus type 1 (HIV-1) is a retrovirus that causes acquired immunodeficiency syndrome (AIDS). HIV-1 Tat protein upregulates transcriptional transactivation. The nucleocapsid protein NC of HIV-1 is a component of virion and plays a key role in genome packaging. Herein, we have demonstrated the interaction between NC and Tat by means of a yeast two-hybrid assay, GST pull-down analysis, co-immunoprecipitation and subcellular colocalization analysis. We observed that the level of Tat was significantly reduced in the presence of NC. But NC did not affect mRNA expression level of Tat. The level of Tat in the presence of NC was increased by treating cells with a proteasome inhibitor, MG132. The ubiquitination state of Tat was not seen to increase in the presence of NC, suggesting the proteasomal degradation was independent of ubiquitination. Lowered level of Tat in the presence of NC led to a decrease in Tat-mediated transcriptional transactivation. Viruses 2013-04-23 5 4 Article 10.3390/v5041143 1143 1152 1999-4915 2013-04-23 doi: 10.3390/v5041143 Hye-Won Hong Seong-Wook Lee Heejoon Myung http://www.mdpi.com/1999-4915/5/4/1131 Latent Membrane Protein 1 (LMP1) is a primary target for controlling tumorigenesis in Epstein-Barr virus related malignancies; in this study, we aimed to develop a specific antibody against the TES1 domain of the oncogenic LMP1. We screened a full human naïve Fab phage library against TES1 peptide, which consisted of C terminal-activating regions proximal 44 amino acids. After three rounds of panning, enrichment and testing by phage ELISA and further analyzed by DNA sequencing, we selected a phage clone with the highest affinity to LMP1-TES1 and designated it as htesFab. The positive clone was expressed in Escherichia coli and the purified htesFab was characterized for its binding specificity and affinity to LMP1. ELISA, immunofluorescence and FACS analysis confirmed that htesFab could recognize LMP1 TES1 both in vitro and in LMP1 expressing HNE2-LMP1 cells. Furthermore, MTT assay showed that htesFab inhibited the proliferation of HNE2-LMP1 cells in a dose-dependent manner. In summary, this study reported the isolation and characterization of human Fab, which specifically targets the C terminal region/TES1 of LMP1, and has potential to be developed as novel tool for the diagnosis and therapy of Epstein-Barr virus related carcinoma Viruses 2013-04-22 5 4 Article 10.3390/v5041131 1131 1142 1999-4915 2013-04-22 doi: 10.3390/v5041131 Dawei Zhang Yuan Mao Qing Cao Lin Xiong Juan Wen Renjie Chen Jin Zhu http://www.mdpi.com/1999-4915/5/4/1114 The mechanism by which the foot-and-mouth disease virus (FMDV) initiates infection of cells is thought to involve the attachment of the viral capsid to host integrins on the surface of target cells. However, the role of integrins in FMDV infection still needs to be fully understood, although it has been demonstrated that integrin αvβ6 interferes with FMDV in vitro and results in neutralization of its infectivity. In the present study, we describe the cloning and sequencing of suckling mouse integrin β6 and the subsequent expression of two segments of integrin β6 extracellular domains: β6-1 (which contains the ligand-binding domain) and β6-2. Sequencing of the mouse integrin β6 subunit revealed close homology (~90%) with its human counterpart. When recombinant integrin extracellular domains β6-1 and β6-2 formulated with adjuvant were inoculated into guinea pigs, anti-integrin antibody expression was high before FMDV challenge. Interestingly, guinea pigs (50%) inoculated with integrin β6-1 were protected from FMDV infection; in contrast, none of the animals inoculated with integrin β6-2 were protected. This result indicates that an integrin blockade may be able to interfere with FMDV infection in vivo, which raises the possibility that targeting integrin in vivo may be the basis for a new strategy to control FMDV infection. Viruses 2013-04-19 5 4 Article 10.3390/v5041114 1114 1130 1999-4915 2013-04-19 doi: 10.3390/v5041114 Yan Zhang Yingjun Sun Fan Yang Jianhong Guo Jijun He Qiong Wu Weijun Cao Lv Lv Haixue Zheng Zhidong Zhang http://www.mdpi.com/1999-4915/5/4/1099 The chrysanthemum (Dendranthema X grandiflorum) belongs to the family Asteraceae and it is one of the most popular flowers in the world. Viroids are the smallest known plant pathogens. They consist of a circular, single-stranded RNA, which does not encode a protein. Chrysanthemums are a common host for two different viroids, the Chrysanthemum stunt viroid (CSVd) and the Chrysanthemum chlorotic mottle viroid (CChMVd). These viroids are quite different from each other in structure and function. Here, we reviewed research associated with CSVd and CChMVd that covered disease symptoms, identification, host range, nucleotide sequences, phylogenetic relationships, structures, replication mechanisms, symptom determinants, detection methods, viroid elimination, and development of viroid resistant chrysanthemums, among other studies. We propose that the chrysanthemum and these two viroids represent convenient genetic resources for host–viroid interaction studies. Viruses 2013-04-17 5 4 Review 10.3390/v5041099 1099 1113 1999-4915 2013-04-17 doi: 10.3390/v5041099 Won Cho Yeonhwa Jo Kyoung-Min Jo Kook-Hyung Kim http://www.mdpi.com/1999-4915/5/4/1075 Like all other viruses, a successful egress of functional particles from infected cells is a prerequisite for foamy virus (FV) spread within the host. The budding process of FVs involves steps, which are shared by other retroviruses, such as interaction of the capsid protein with components of cellular vacuolar protein sorting (Vps) machinery via late domains identified in some FV capsid proteins. Additionally, there are features of the FV budding strategy quite unique to the spumaretroviruses. This includes secretion of non-infectious subviral particles and a strict dependence on capsid-glycoprotein interaction for release of infectious virions from the cells. Virus-like particle release is not possible since FV capsid proteins lack a membrane-targeting signal. It is noteworthy that in experimental systems, the important capsid-glycoprotein interaction could be bypassed by fusing heterologous membrane-targeting signals to the capsid protein, thus enabling glycoprotein-independent egress. Aside from that, other systems have been developed to enable envelopment of FV capsids by heterologous Env proteins. In this review article, we will summarize the current knowledge on FV budding, the viral components and their domains involved as well as alternative and artificial ways to promote budding of FV particle structures, a feature important for alteration of target tissue tropism of FV-based gene transfer systems. Viruses 2013-04-10 5 4 Review 10.3390/v5041075 1075 1098 1999-4915 2013-04-10 doi: 10.3390/v5041075 Sylvia Hütter Irena Zurnic Dirk Lindemann http://www.mdpi.com/1999-4915/5/4/1055 Here we review viral and cellular requirements for entry and intracellular trafficking of foamy viruses (FVs) resulting in integration of viral sequences into the host cell genome. The virus encoded glycoprotein harbors all essential viral determinants, which are involved in absorption to the host membrane and triggering the uptake of virus particles. However, only recently light was shed on some details of FV’s interaction with its host cell receptor(s). Latest studies indicate glycosaminoglycans of cellular proteoglycans, particularly heparan sulfate, to be of utmost importance. In a species-specific manner FVs encounter endogenous machineries of the target cell, which are in some cases exploited for fusion and further egress into the cytosol. Mostly triggered by pH-dependent endocytosis, viral and cellular membranes fuse and release naked FV capsids into the cytoplasm. Intact FV capsids are then shuttled along microtubules and are found to accumulate nearby the centrosome where they can remain in a latent state for extended time periods. Depending on the host cell cycle status, FV capsids finally disassemble and, by still poorly characterized mechanisms, the preintegration complex gets access to the host cell chromatin. Host cell mitosis finally allows for viral genome integration, ultimately starting a new round of viral replication. Viruses 2013-04-08 5 4 Review 10.3390/v5041055 1055 1074 1999-4915 2013-04-08 doi: 10.3390/v5041055 Ursula Berka Martin Hamann Dirk Lindemann http://www.mdpi.com/1999-4915/5/4/1042 The Epstein-Barr virus (EBV) double-stranded DNA genome is subject to extensive epigenetic regulation. Large consortiums and individual labs have generated a vast number of genome-wide data sets on human lymphoblastoid and other cell lines latently infected with EBV. Analysis of these data sets reveals important new information on the properties of the host and viral chromosome structure organization and epigenetic modifications. We discuss the mapping of these data sets and the subsequent insights into the chromatin structure and transcription factor binding patterns on latent EBV genomes. Colocalization of multiple histone modifications and transcription factors at regulatory loci are considered in the context of the biology and regulation of EBV. Viruses 2013-04-02 5 4 Commentary 10.3390/v5041042 1042 1054 1999-4915 2013-04-02 doi: 10.3390/v5041042 Aaron Arvey Italo Tempera Paul Lieberman http://www.mdpi.com/1999-4915/5/4/1023 Gag proteins play an important role in many stages of the retroviral replication cycle. They orchestrate viral assembly, interact with numerous host cell proteins, engage in regulation of viral gene expression, and provide the main driving force for virus intracellular trafficking and budding. Foamy Viruses (FV), also known as spumaviruses, display a number of unique features among retroviruses. Many of these features can be attributed to their Gag proteins. FV Gag proteins lack characteristic orthoretroviral domains like membrane-binding domains (M domains), the major homology region (MHR), and the hallmark Cys-His motifs. In contrast, they contain several distinct domains such as the essential Gag-Env interaction domain and the glycine and arginine rich boxes (GR boxes). Furthermore, FV Gag only undergoes limited maturation and follows an unusual pathway for nuclear translocation. This review summarizes the known FV Gag domains and motifs and their functions. In particular, it provides an overview of the unique structural and functional properties that distinguish FV Gag proteins from orthoretroviral Gag proteins. Viruses 2013-03-26 5 4 Review 10.3390/v5041023 1023 1041 1999-4915 2013-03-26 doi: 10.3390/v5041023 Erik Müllers http://www.mdpi.com/1999-4915/5/3/998 The development of high-throughput technologies allows for evaluating gene expression at the whole-genome level. Together with proteomic and metabolomic studies, these analyses have resulted in the identification of plant genes whose function or expression is altered as a consequence of pathogen attacks. Members of the Tomato yellow leaf curl virus (TYLCV) complex are among the most important pathogens impairing production of agricultural crops worldwide. To understand how these geminiviruses subjugate plant defenses, and to devise counter-measures, it is essential to identify the host genes affected by infection and to determine their role in susceptible and resistant plants. We have used a reverse genetics approach based on Tobacco rattle virus-induced gene silencing (TRV-VIGS) to uncover genes involved in viral infection of susceptible plants, and to identify genes underlying virus resistance. To identify host genes with a role in geminivirus infection, we have engineered a Nicotiana benthamiana line, coined 2IRGFP, which over-expresses GFP upon virus infection. With this system, we have achieved an accurate description of the dynamics of virus replication in space and time. Upon silencing selected N. benthamiana genes previously shown to be related to host response to geminivirus infection, we have identified eighteen genes involved in a wide array of cellular processes. Plant genes involved in geminivirus resistance were studied by comparing two tomato lines: one resistant (R), the other susceptible (S) to the virus. Sixty-nine genes preferentially expressed in R tomatoes were identified by screening cDNA libraries from infected and uninfected R and S genotypes. Out of the 25 genes studied so far, the silencing of five led to the total collapse of resistance, suggesting their involvement in the resistance gene network. This review of our results indicates that TRV-VIGS is an exquisite reverse genetics tool that may provide new insights into the molecular mechanisms underlying plant infection and resistance to infection by begomoviruses. Viruses 2013-03-22 5 3 Review 10.3390/v5030998 998 1022 1999-4915 2013-03-22 doi: 10.3390/v5030998 Henryk Czosnek Assaf Eybishtz Dagan Sade Rena Gorovits Iris Sobol Eduardo Bejarano Tábata Rosas-Díaz Rosa Lozano-Durán http://www.mdpi.com/1999-4915/5/3/981 DEB025 (alisporivir) is a synthetic cyclosporine with inhibitory activity against human immunodeficiency virus type-1 (HIV-1) and hepatitis C virus (HCV). It binds to cyclophilin A (CypA) and blocks essential functions of CypA in the viral replication cycles of both viruses. DEB025 inhibits clinical HIV-1 isolates in vitro and decreases HIV-1 virus load in the majority of patients. HIV-1 isolates being naturally resistant to DEB025 have been detected in vitro and in nonresponder patients. By sequence analysis of their capsid protein (CA) region, two amino acid polymorphisms that correlated with DEB025 resistance were identified: H87Q and I91N, both located in the CypA-binding loop of the CA protein of HIV-1. The H87Q change was by far more abundant than I91N. Additional polymorphisms in the CypA-binding loop (positions 86, 91 and 96), as well as in the N-terminal loop of CA were detected in resistant isolates and are assumed to contribute to the degree of resistance. These amino acid changes may modulate the conformation of the CypA-binding loop of CA in such a way that binding and/or isomerase function of CypA are no longer necessary for virus replication. The resistant HIV-1 isolates thus are CypA-independent. Viruses 2013-03-22 5 3 Article 10.3390/v5030981 981 997 1999-4915 2013-03-22 doi: 10.3390/v5030981 Philippe Gallay Roger Ptak Michael Bobardt Jean-Maurice Dumont Grégoire Vuagniaux Brigitte Rosenwirth http://www.mdpi.com/1999-4915/5/3/954 Chronic hepatitis C is characterized by metabolic disorders and a microenvironment in the liver dominated by oxidative stress, inflammation and regeneration processes that lead in the long term to hepatocellular carcinoma. Many lines of evidence suggest that mitochondrial dysfunctions, including modification of metabolic fluxes, generation and elimination of oxidative stress, Ca2+ signaling and apoptosis, play a central role in these processes. However, how these dysfunctions are induced by the virus and whether they play a role in disease progression and neoplastic transformation remains to be determined. Most in vitro studies performed so far have shown that several of the hepatitis C virus (HCV) proteins localize to mitochondria, but the consequences of these interactions on mitochondrial functions remain contradictory, probably due to the use of artificial expression and replication systems. In vivo studies are hampered by the fact that innate and adaptive immune responses will overlay mitochondrial dysfunctions induced directly in the hepatocyte by HCV. Thus, the molecular aspects underlying HCV-induced mitochondrial dysfunctions and their roles in viral replication and the associated pathology need yet to be confirmed in the context of productively replicating virus and physiologically relevant in vitro and in vivo model systems. Viruses 2013-03-21 5 3 Review 10.3390/v5030954 954 980 1999-4915 2013-03-21 doi: 10.3390/v5030954 Charlène Brault Pierre Levy Birke Bartosch http://www.mdpi.com/1999-4915/5/3/928 Vaccinia virus (VACV) was used as a surrogate of variola virus (VARV) (genus Orthopoxvirus), the causative agent of smallpox, to study Orthopoxvirus infection. VARV is principally transmitted between humans by aerosol droplets. Once inhaled, VARV first infects the respiratory tract where it could encounter surfactant components, such as soluble pattern recognition receptors. Surfactant protein D (SP-D), constitutively present in the lining fluids of the respiratory tract, plays important roles in innate host defense against virus infection. We investigated the role of SP-D in VACV infection and studied the A27 viral protein involvement in the interaction with SP-D. Interaction between SP-D and VACV caused viral inhibition in a lung cell model. Interaction of SP-D with VACV was mediated by the A27 viral protein. Binding required Ca2+ and interactions were blocked in the presence of excess of SP-D saccharide ligands. A27, which lacks glycosylation, directly interacted with SP-D. The interaction between SP-D and the viral particle was also observed using electron microscopy. Infection of mice lacking SP-D (SP-D-/-) resulted in increased mortality compared to SP-D+/+ mice. Altogether, our data show that SP-D participates in host defense against the vaccinia virus infection and that the interaction occurs with the viral surface protein A27. Viruses 2013-03-21 5 3 Article 10.3390/v5030928 928 953 1999-4915 2013-03-21 doi: 10.3390/v5030928 Julien Perino Nicole Thielens Erika Crouch Danièle Spehner Jean-Marc Crance Anne-Laure Favier http://www.mdpi.com/1999-4915/5/3/902 Highly active antiretroviral therapy (HAART) has limited the replication and spread of the human immunodeficiency virus (HIV). However, despite treatment, HIV infection persists in latently infected reservoirs, and once therapy is interrupted, viral replication rebounds quickly. Extensive efforts are being directed at eliminating these cell reservoirs. This feat can be achieved by reactivating latent HIV while administering drugs that prevent new rounds of infection and allow the immune system to clear the virus. However, current approaches to HIV eradication have not been effective. Moreover, as HIV latency is multifactorial, the significance of each of its molecular mechanisms is still under debate. Among these, transcriptional repression as a result of reduced levels and activity of the positive transcription elongation factor b (P-TEFb: CDK9/cyclin T) plays a significant role. Therefore, increasing levels of P-TEFb expression and activity is an excellent strategy to stimulate viral gene expression. This review summarizes the multiple steps that cause HIV to enter into latency. It positions the interplay between transcriptionally active and inactive host transcriptional activators and their viral partner Tat as valid targets for the development of new strategies to reactivate latent viral gene expression and eradicate HIV. Viruses 2013-03-21 5 3 Article 10.3390/v5030902 902 927 1999-4915 2013-03-21 doi: 10.3390/v5030902 Ran Taube Matija Peterlin http://www.mdpi.com/1999-4915/5/3/901 A funding designation in our 2012 Viruses publication, doi: 10.3390/v4101844 (Viruses 2012, 4, 1844-1864), contained an incorrect digit. On page 1860, (Acknowledgments), U19 support was incorrectly listed as AI076113. The correct designation is AI096113. Viruses 2013-03-20 5 3 Correction 10.3390/v5030901 901 901 1999-4915 2013-03-20 doi: 10.3390/v5030901 Jeanne Sisk Janice Clements Kenneth Witwer http://www.mdpi.com/1999-4915/5/3/886 Foamy viruses (FVs) differ from all other genera of retroviruses (orthoretroviruses) in many aspects of viral replication. In this review, we discuss FV assembly, with special emphasis on Pol incorporation. FV assembly takes place intracellularly, near the pericentriolar region, at a site similar to that used by betaretroviruses. The regions of Gag, Pol and genomic RNA required for viral assembly are described. In contrast to orthoretroviral Pol, which is synthesized as a Gag-Pol fusion protein and packaged through Gag-Gag interactions, FV Pol is synthesized from a spliced mRNA lacking all Gag sequences. Thus, encapsidation of FV Pol requires a different mechanism. We detail how WT Pol lacking Gag sequences is incorporated into virus particles. In addition, a mutant in which Pol is expressed as an orthoretroviral-like Gag-Pol fusion protein is discussed. We also discuss temporal regulation of the protease, reverse transcriptase and integrase activities of WT FV Pol. Viruses 2013-03-20 5 3 Review 10.3390/v5030886 886 900 1999-4915 2013-03-20 doi: 10.3390/v5030886 Eun-Gyung Lee Carolyn Stenbak Maxine Linial http://www.mdpi.com/1999-4915/5/3/873 In this review we propose a partially hypothetical model of respiratory syncytial virus (RSV) binding and entry to the cell that includes the recently discovered RSV receptor nucleolin, in an attempt to stimulate further inquiry in this research area. RSV binding and entry is likely to be a two-step process, the first involving the attachment of the virus to the cell membrane, which may be enhanced by electrostatic interactions with cellular glycoproteins/heparin and the viral G protein, and the second involving fusion to the cell membrane mediated by the viral F protein and a specific cellular fusion receptor. With our recent discovery of nucleolin as a functional fusion receptor for RSV, comes the possibility of a number of new approaches to the development of novel strategies for RSV prophylaxis and therapy, as well as raising some new questions concerning the pathobiology of RSV infection and tropism. Viruses 2013-03-19 5 3 Review 10.3390/v5030873 873 885 1999-4915 2013-03-19 doi: 10.3390/v5030873 Peter Mastrangelo Richard Hegele http://www.mdpi.com/1999-4915/5/3/858 This review focuses on the significance of deregulation of epigenetic mechanisms by the hepatitis B virus (HBV) X protein in hepatocarcinogenesis and HBV replication. Epigenetic mechanisms, DNA methylation, and specific histone modifications, e.g., trimethylation of H3 on lysine-27 or lysine-4, maintain ‘cellular memory’ by silencing expression of lineage-inducing factors in stem cells and conversely, of pluripotency factors in differentiated cells. The X protein has been reported to induce expression of DNA methyltransferases (DNMTs), likely promoting epigenetic changes during hepatocarcinogenesis. Furthermore, in cellular and animal models of X-mediated oncogenic transformation, protein levels of chromatin modifying proteins Suz12 and Znf198 are down-regulated. Suz12 is essential for the Polycomb Repressive Complex 2 (PRC2) mediating the repressive trimethylation of H3 on lysine-27 (H3K27me3). Znf198, stabilizes the LSD1-CoREST-HDAC complex that removes, via lysine demethylase1 (LSD1), the activating trimethylation of H3 on lysine-4 (H3K4me3). Down-regulation of Suz12 also occurs in liver tumors of woodchucks chronically infected by woodchuck hepatitis virus, an animal model recapitulating HBV-mediated hepatocarcinogenesis in humans. Significantly, subgroups of HBV-induced liver cancer re-express hepatoblast and fetal markers, and imprinted genes, suggesting hepatocyte reprogramming during oncogenic transformation. Lastly, down-regulation of Suz12 and Znf198 enhances HBV replication. Collectively, these observations suggest deregulation of epigenetic mechanisms by HBV X protein influences both the viral cycle and the host cell. Viruses 2013-03-18 5 3 Review 10.3390/v5030858 858 872 1999-4915 2013-03-18 doi: 10.3390/v5030858 Ourania Andrisani http://www.mdpi.com/1999-4915/5/3/834 The central nervous system (CNS) harbors highly differentiated cells, such as neurons that are essential to coordinate the functions of complex organisms. This organ is partly protected by the blood-brain barrier (BBB) from toxic substances and pathogens carried in the bloodstream. Yet, neurotropic viruses can reach the CNS either by crossing the BBB after viremia, or by exploiting motile infected cells as Trojan horses, or by using axonal transport. Type I and type III interferons (IFNs) are cytokines that are critical to control early steps of viral infections. Deficiencies in the IFN pathway have been associated with fatal viral encephalitis both in humans and mice. Therefore, the IFN system provides an essential protection of the CNS against viral infections. Yet, basal activity of the IFN system appears to be low within the CNS, likely owing to the toxicity of IFN to this organ. Moreover, after viral infection, neurons and oligodendrocytes were reported to be relatively poor IFN producers and appear to keep some susceptibility to neurotropic viruses, even in the presence of IFN. This review addresses some trends and recent developments concerning the role of type I and type III IFNs in: i) preventing neuroinvasion and infection of CNS cells; ii) the identity of IFN-producing cells in the CNS; iii) the antiviral activity of ISGs; and iv) the activity of viral proteins of neurotropic viruses that target the IFN pathway. Viruses 2013-03-15 5 3 Review 10.3390/v5030834 834 857 1999-4915 2013-03-15 doi: 10.3390/v5030834 Frédéric Sorgeloos Marguerite Kreit Pascale Hermant Cécile Lardinois Thomas Michiels http://www.mdpi.com/1999-4915/5/3/824 While PCR amplicons extend to a few thousand bases, the length of sequences from direct Sanger sequencing is limited to 500–800 nucleotides. Therefore, several fragments may be required to cover an amplicon, a gene or an entire genome. These fragments are typically sequenced in an overlapping fashion and assembled by manually sliding and aligning the sequences visually. This is time-consuming, repetitive and error-prone, and further complicated by circular genomes. An online tool merging two to twelve long overlapping sequence fragments was developed. Either chromatograms or FASTA files are submitted to the tool, which trims poor quality ends of chromatograms according to user-specified parameters. Fragments are assembled into a single sequence by repeatedly calling the EMBOSS merger tool in a consecutive manner. Output includes the number of trimmed nucleotides, details of each merge, and an optional alignment to a reference sequence. The final merge sequence is displayed and can be downloaded in FASTA format. All output files can be downloaded as a ZIP archive. This tool allows for easy and automated assembly of overlapping sequences and is aimed at researchers without specialist computer skills. The tool is genome- and organism-agnostic and has been developed using hepatitis B virus sequence data. Viruses 2013-03-12 5 3 Article 10.3390/v5030824 824 833 1999-4915 2013-03-12 doi: 10.3390/v5030824 Trevor Bell Anna Kramvis http://www.mdpi.com/1999-4915/5/3/806 Studying the coevolutionary dynamics between bacteria and the bacteriophage viruses that infect them is critical to understanding both microbial diversity and ecosystem functioning. Phages can play a key role in shaping bacterial population dynamics and can significantly alter both intra- and inter-specific competition among bacterial hosts. Predicting how phages might influence community stability and apparent competition, however, requires an understanding of how bacteria-phage interaction networks evolve as a function of host diversity and community dynamics. Here, we first review the progress that has been made in understanding phage specificity, including the use of experimental evolution, we then introduce a new dataset on natural bacteriophages collected from the phyllosphere of horse chestnut trees, and finally we highlight that bacterial sensitivity to phage is rarely a binary trait and that this variation should be taken into account and reported. We emphasize that there is currently insufficient evidence to make broad generalizations about phage host range in natural populations, the limits of phage adaptation to novel hosts, or the implications of phage specificity in shaping microbial communities. However, the combination of experimental and genomic approaches with the study of natural communities will allow new insight to the evolution and impact of phage specificity within complex bacterial communities. Viruses 2013-03-11 5 3 Review 10.3390/v5030806 806 823 1999-4915 2013-03-11 doi: 10.3390/v5030806 Britt Koskella Sean Meaden http://www.mdpi.com/1999-4915/5/3/792 The NSP4 protein is a multifunctional protein that plays a role in the morphogenesis and pathogenesis of the rotavirus. Although NSP4 is considered an enterotoxin, the relationship between gastroenteritis severity and amino acid variations in NSP4 of the human rotavirus remains unclear. In this study, we analyzed the sequence diversity of NSP4 and the severity of gastroenteritis of children with moderate to severe gastroenteritis. The rotavirus-infected children were hospitalized before the rotavirus vaccine program in Mexico. All children had diarrhea within 1-4 days, 44 (88%) were vomiting and 35 (70%) had fevers. The severity analysis showed that 13 (26%) cases had mild gastroenteritis, 23 (46%) moderate gastroenteritis and 14 (28%) severe. NSP4 phylogenetic analysis showed three clusters within the genotype E1. Sequence analysis revealed similar mutations inside each cluster, and an uncommon variation in residue 144 was found in five of the Mexican NSP4 sequences. Most of the amino acid variations were located in the VP4 and VP6 binding site domains, with no relationship to different grades of gastroenteritis. This finding indicates that severe gastroenteritis caused by the rotavirus appears to be related to diverse viral or cellular factors instead of NSP4 activity as a unique pathogenic factor. Viruses 2013-03-11 5 3 Article 10.3390/v5030792 792 805 1999-4915 2013-03-11 doi: 10.3390/v5030792 Guadalupe González-Ochoa Griselda Menchaca Carlos Hernández Cristina Rodríguez Reyes Tamez Juan Contreras http://www.mdpi.com/1999-4915/5/3/777 Infection by respiratory syncytial virus (RSV) affects approximately 33 million infants annually worldwide and is a major cause of hospitalizations. Helper T lymphocytes (Th) play a central role in the immune response during such infections. However, Th lymphocytes that produce interleukin 17 (IL-17), known as Th17 lymphocytes, in addition to been protective can also cause pathology that accompany this type of infection. The protective effects of Th17 is associated with better prognosis in most infected individuals but heightened Th17 responses causes inflammation and pathology in others. Studies employing animal models haves shown that activated Th17 lymphocytes recruit neutrophils and facilitate tertiary lymphoid structure development in infected lungs. However, IL-17 also inhibits the ability of CD8+ lymphocytes to clear viral particles and acts synergistically with the innate immune system to exacerbate inflammation. Furthermore, IL-17 enhances IL-13 production which, in turn, promotes the activation of Th2 lymphocytes and excessive mucus production. Studies of these animal models have also shown that a lack of, or inadequate, responses by the Th1 subset of T lymphocytes enhances Th17-mediated responses and that this is detrimental during RSV co-infection in experimental asthma. The available evidence, therefore, indicates that Th17 can play contradictory roles during RSV infections. The factors that determine the shift in the balance between beneficial and adverse Th17 mediated effects during RSV infection remains to be determined. Viruses 2013-03-05 5 3 Review 10.3390/v5030777 777 791 1999-4915 2013-03-05 doi: 10.3390/v5030777 Jonas Bystrom Nasra Al-Adhoubi Mohammed Al-Bogami Ali Jawad Rizgar Mageed http://www.mdpi.com/1999-4915/5/2/758 Banana bunchy top virus (BBTV) is the most destructive pathogenic virus of banana plants worldwide. The virus is transmitted in a circulative non-propagative manner by the banana aphid, Pentalonia nigronervosa Coquerel. In this work, we examined the localization, accumulation, and transmission efficiency of BBTV in four laboratory-established lineages of Pentalonia aphids derived from four different host plants: taro (Colocasia esculenta), heliconia (Heliconia spp.), red ginger (Alpinia purpurata), and banana (Musa sp.). Mitochondrial sequencing identified three and one lineages as Pentalonia caladii van der Goot, a recently proposed species, and P. nigronervosa, respectively. Microsatellite analysis separated the aphid lineages into four distinct genotypes. The transmission of BBTV was tested using leaf disk and whole-plant assays, both of which showed that all four lineages are competent vectors of BBTV, although the P. caladii from heliconia transmitted BBTV to the leaf disks at a significantly lower rate than did P. nigronervosa. The concentration of BBTV in dissected guts, haemolymph, and salivary glands was quantified by real-time PCR. The BBTV titer reached similar concentrations in the guts, haemolymph, and salivary glands of aphids from all four lineages tested. Furthermore, immunofluorescence assays showed that BBTV antigens localized to the anterior midguts and the principal salivary glands, demonstrating a similar pattern of translocations across the four lineages. The results reported in this study showed for the first time that P. caladii is a competent vector of BBTV. Viruses 2013-02-22 5 2 Article 10.3390/v5020758 758 776 1999-4915 2013-02-22 doi: 10.3390/v5020758 Shizu Watanabe April Greenwell Alberto Bressan http://www.mdpi.com/1999-4915/5/2/753 The SupT1 cell line supports optimal HIV-1 replication, and prolonged in vitro replication in SupT1 cells renders the virus significantly less virulent. This raises the question of whether the infusion of SupT1 cells could be used as a cell-based therapy to induce a pacific coexistence between the HIV virus and its human host. In a recent study, I investigated this potential therapeutic strategy in vitro. The results suggested that this approach should be further explored in HIV-susceptible animal models. Such studies may lead to the development of a functional cure for HIV infection. Viruses 2013-02-21 5 2 Opinion 10.3390/v5020753 753 757 1999-4915 2013-02-21 doi: 10.3390/v5020753 Jonathan Fior http://www.mdpi.com/1999-4915/5/2/732 The nervous system is the target for acute encephalitic viral infections, as well as a reservoir for persisting viruses. Intrathecal antibody (Ab) synthesis is well documented in humans afflicted by infections associated with neurological complications, as well as the demyelinating disease, multiple sclerosis. This review focuses on the origin, recruitment, maintenance, and biological relevance of Ab-secreting cells (ASC) found in the central nervous system (CNS) following experimental neurotropic RNA virus infections. We will summarize evidence for a highly dynamic, evolving humoral response characterized by temporal alterations in B cell subsets, proliferation, and differentiation. Overall local Ab plays a beneficial role via complement-independent control of virus replication, although cross or self-reactive Ab to CNS antigens may contribute to immune-mediated pathogenesis during some infections. Importantly, protective Ab exert anti-viral activity not only by direct neutralization, but also by binding to cell surface-expressed viral glycoproteins. Ab engagement of viral glycoproteins blocks budding and mediates intracellular signaling leading to restored homeostatic and innate functions. The sustained Ab production by local ASC, as well as chemokines and cytokines associated with ASC recruitment and retention, are highlighted as critical components of immune control. Viruses 2013-02-15 5 2 Review 10.3390/v5020732 732 752 1999-4915 2013-02-15 doi: 10.3390/v5020732 Timothy Phares Stephen Stohlman Cornelia Bergmann http://www.mdpi.com/1999-4915/5/2/708 Extensive experimental work has conclusively demonstrated that infection with certain types of human papillomaviruses, the so-called high-risk human papillomavirus (HR-HPV), represent a most powerful human carcinogen. However, neoplastic growth is a rare and inappropriate outcome in the natural history of HPV, and a number of other events have to concur in order to induce the viral infection into the (very rare) neoplastic transformation. From this perspective, a number of putative viral, host, and environmental co-factors have been proposed as potential candidates. Among them oxidative stress (OS) is an interesting candidate, yet comparatively underexplored. OS is a constant threat to aerobic organisms being generated during mitochondrial oxidative phosphorylation, as well as during inflammation, infections, ionizing irradiation, UV exposure, mechanical and chemical stresses. Epithelial tissues, the elective target for HPV infection, are heavily exposed to all named sources of OS. Two different types of cooperative mechanisms are presumed to occur between OS and HPV: I) The OS genotoxic activity and the HPV-induced genomic instability concur independently to the generation of the molecular damage necessary for the emergence of neoplastic clones. This first mode is merely a particular form of co-carcinogenesis; and II) OS specifically interacts with one or more molecular stages of neoplastic initiation and/or progression induced by the HPV infection. This manuscript was designed to summarize available data on this latter hypothesis. Experimental data and indirect evidences on promoting the activity of OS in viral infection and viral integration will be reviewed. The anti-apoptotic and pro-angiogenetic role of NO (nitric oxide) and iNOS (inducible nitric oxide synthase) will be discussed together with the OS/HPV cooperation in inducing cancer metabolism adaptation. Unexplored/underexplored aspects of the OS interplay with the HPV-driven carcinogenesis will be highlighted. The aim of this paper is to stimulate new areas of study and innovative approaches. Viruses 2013-02-12 5 2 Review 10.3390/v5020708 708 731 1999-4915 2013-02-12 doi: 10.3390/v5020708 Federico De Marco http://www.mdpi.com/1999-4915/5/2/678 Alphaherpesviruses are pathogens that invade the nervous systems of their mammalian hosts. Directional spread of infection in the nervous system is a key component of the viral lifecycle and is critical for the onset of alphaherpesvirus-related diseases. Many alphaherpesvirus infections originate at peripheral sites, such as epithelial tissues, and then enter neurons of the peripheral nervous system (PNS), where lifelong latency is established. Following reactivation from latency and assembly of new viral particles, the infection typically spreads back out towards the periphery. These spread events result in the characteristic lesions (cold sores) commonly associated with herpes simplex virus (HSV) and herpes zoster (shingles) associated with varicella zoster virus (VZV). Occasionally, the infection spreads transsynaptically from the PNS into higher order neurons of the central nervous system (CNS). Spread of infection into the CNS, while rarer in natural hosts, often results in severe consequences, including death. In this review, we discuss the viral and cellular mechanisms that govern directional spread of infection in the nervous system. We focus on the molecular events that mediate long distance directional transport of viral particles in neurons during entry and egress. Viruses 2013-02-11 5 2 Review 10.3390/v5020678 678 707 1999-4915 2013-02-11 doi: 10.3390/v5020678 Tal Kramer Lynn Enquist http://www.mdpi.com/1999-4915/5/2/663 This work describes peptides from non-structural proteins (nsp) of porcine reproductive and respiratory syndrome virus (PRRSV) predicted as potential T cell epitopes by bioinfornatics and tested for their ability to induce IFN-γ and IL-10 responses. Pigs immunized with either genotype 1 or genotype 2 PRRSV attenuated vaccines (n=5/group) and unvaccinated pigs (n = 4) were used to test the peptides. Swine leukocyte antigen haplotype of each pig was also determined. Pigs were initially screened for IFN-γ responses (ELISPOT) and three peptides were identified; two of them in non-conserved segments of nsp2 and nsp5 and the other in a conserved region of nsp5 peptide. Then, peptides were screened for IL-10 inducing properties. Six peptides were found to induce IL-10 release in PBMC and some of them were also able to inhibit IFN-γ responses on PHA-stimulated cells. Interestingly, the IFN-γ low responder pigs against PRRSV were mostly homozygous for their SLA haplotypes. In conclusion, these results indicate that nsp of PRRSV contain T-cell epitopes inducing IFN-γ responses as well as IL-10 inducing segments with inhibitory capabilities. Viruses 2013-02-11 5 2 Article 10.3390/v5020663 663 677 1999-4915 2013-02-11 doi: 10.3390/v5020663 Alexel Burgara-Estrella Ivan Díaz Irene Rodríguez-Gómez Sabine Essler Jesús Hernández Enric Mateu http://www.mdpi.com/1999-4915/5/2/654 Accumulation of aggregates rich in an abnormally folded form of the prion protein characterize the neurodegeneration caused by transmissible spongiform encephalopathies (TSEs). The molecular triggers of plaque formation and neurodegeneration remain unknown, but analyses of TSE-infected brain homogenates and preparations enriched for abnormal prion protein suggest that reduced levels of copper and increased levels of manganese are associated with disease. The objectives of this study were to: (1) assess copper and manganese levels in healthy and TSE-infected Syrian hamster brain homogenates; (2) determine if the distribution of these metals can be mapped in TSE-infected brain tissue using X-ray photoelectron emission microscopy (X-PEEM) with synchrotron radiation; and (3) use X-PEEM to assess the relative amounts of copper and manganese in prion plaques in situ. In agreement with studies of other TSEs and species, we found reduced brain levels of copper and increased levels of manganese associated with disease in our hamster model. We also found that the in situ levels of these metals in brainstem were sufficient to image by X-PEEM. Using immunolabeled prion plaques in directly adjacent tissue sections to identify regions to image by X-PEEM, we found a statistically significant relationship of copper-manganese dysregulation in prion plaques: copper was depleted whereas manganese was enriched. These data provide evidence for prion plaques altering local transition metal distribution in the TSE-infected central nervous system. Viruses 2013-02-11 5 2 Communication 10.3390/v5020654 654 662 1999-4915 2013-02-11 doi: 10.3390/v5020654 Christopher Johnson P.U.P.A. Gilbert Mike Abrecht Katherine Baldwin Robin Russell Joel Pedersen Judd Aiken Debbie McKenzie http://www.mdpi.com/1999-4915/5/2/619 The initiation of bacteriophage λ replication depends upon interactions between the oriλ DNA site, phage proteins O and P, and E. coli host replication proteins. P exhibits a high affinity for DnaB, the major replicative helicase for unwinding double stranded DNA. The concept of P-lethality relates to the hypothesis that P can sequester DnaB and in turn prevent cellular replication initiation from oriC. Alternatively, it was suggested that P-lethality does not involve an interaction between P and DnaB, but is targeted to DnaA. P-lethality is assessed by examining host cells for transformation by ColE1-type plasmids that can express P, and the absence of transformants is attributed to a lethal effect of P expression. The plasmid we employed enabled conditional expression of P, where under permissive conditions, cells were efficiently transformed. We observed that ColE1 replication and plasmid establishment upon transformation is extremely sensitive to P, and distinguish this effect from P-lethality directed to cells. We show that alleles of dnaB protect the variant cells from P expression. P-dependent cellular filamentation arose in ΔrecA or lexA[Ind-] cells, defective for SOS induction. Replication propagation and restart could represent additional targets for P interference of E. coli replication, beyond the oriC-dependent initiation step. Viruses 2013-02-06 5 2 Article 10.3390/v5020619 619 653 1999-4915 2013-02-06 doi: 10.3390/v5020619 Sidney Hayes Craig Erker Monique Horbay Kristen Marciniuk Wen Wang Connie Hayes http://www.mdpi.com/1999-4915/5/2/605 Dengue virus (DENV), which is transmitted by Aedes mosquitoes, causes fever and hemorrhagic disorders in humans. The virus entry process mediated through host receptor molecule(s) is crucial for virus propagation and the pathological progression of dengue disease. Therefore, elucidation of the molecular mechanisms underlying virus entry is essential for an understanding of dengue pathology and for the development of effective new anti-dengue agents. DENV binds to its receptor molecules mediated through a viral envelope (E) protein, followed by incorporation of the virus-receptor complex inside cells. The fusion between incorporated virus particles and host endosome membrane under acidic conditions is mediated through the function of DENV E protein. Carbohydrate molecules, such as sulfated glycosaminoglycans (GAG) and glycosphingolipids, and carbohydrate-recognition proteins, termed lectins, inhibit virus entry. This review focuses on carbohydrate-derived entry inhibitors, and also introduces functionally related compounds with similar inhibitory mechanisms against DENV entry. Viruses 2013-02-06 5 2 Review 10.3390/v5020605 605 618 1999-4915 2013-02-06 doi: 10.3390/v5020605 Kazuya Hidari Tomoko Abe Takashi Suzuki http://www.mdpi.com/1999-4915/5/2/595 Entry of HIV-1 into a host cell is a multi-step process, with the viral envelope gp120 and gp41 acting sequentially to mediate the viral attachment, CD4 binding, coreceptor binding, and fusion of the viral and host membranes. The emerging class of antiretroviral agents, collectively known as entry inhibitors, interfere in some of these steps. However, viral diversity has implications for possible differential responses to entry inhibitors, since envelope is the most variable of all HIV genes. Different HIV genetic forms carry in their genomes genetic signatures and polymorphisms that could alter the structure of viral proteins which are targeted by drugs, thus impairing antiretroviral binding and efficacy. This review will examine current research that describes subtype differences in envelope at the genetic level and the effects of mutations on the efficacy of current entry inhibitors. Viruses 2013-02-06 5 2 Review 10.3390/v5020595 595 604 1999-4915 2013-02-06 doi: 10.3390/v5020595 Leonardo Araújo Sabrina Almeida http://www.mdpi.com/1999-4915/5/2/577 Respiratory syncytial virus (RSV) is the etiological agent for a serious lower respiratory tract disease responsible for close to 200,000 annual deaths worldwide. The first infection is generally most severe, while re-infections usually associate with a milder disease. This observation and the finding that re-infection risks are inversely associated with neutralizing antibody titers suggest that immune responses generated toward a first RSV exposure can significantly reduce morbidity and mortality throughout life. For more than half a century, researchers have endeavored to design a vaccine for RSV that can mimic or improve upon natural protective immunity without adverse events. The virus is herein described together with the hurdles that must be overcome to develop a vaccine and some current vaccine development approaches. Viruses 2013-02-05 5 2 Review 10.3390/v5020577 577 594 1999-4915 2013-02-05 doi: 10.3390/v5020577 Rajeev Rudraraju Bart Jones Robert Sealy Sherri Surman Julia Hurwitz http://www.mdpi.com/1999-4915/5/2/568 Cyprinid herpes virus 3 (CyHV-3) diseases have been reported around the world and are associated with high mortalities of koi (Cyprinus carpio). Although little work has been conducted on the molecular analysis of this virus, glycoprotein genes identified in the present study seem to be valuable targets for genetic comparison of this virus. Three envelope glycoprotein genes (ORF25, 65 and 116) of the CyHV-3 isolates from the USA, Israel, Japan and Korea were compared, and interestingly, sequence insertions or deletions were observed in these target regions. In addition, polymorphisms were presented in microsatellite zones from two glycoprotein genes (ORF65 and 116). In phylogenetic tree analysis, the Korean isolate was remarkably distinguished from USA, Israel, Japan isolates. These findings may be suitable for many applications including isolates differentiation and phylogeny studies. Viruses 2013-01-31 5 2 Article 10.3390/v5020568 568 576 1999-4915 2013-01-31 doi: 10.3390/v5020568 Jee Han Ji Kim Tristan Renault Casiano Jr. Sang Shin Jin Jun Se Park http://www.mdpi.com/1999-4915/5/2/550 The nucleotide sequences of a region including S1, S2, 3a, 3b and E genes of twenty-seven infectious bronchitis virus (IBV) isolates in Korea between 1990–2011 were determined and phylogenetic and computational recombination analyses were conducted. The sizes of coding regions of some genes varied among IBV isolates due to deletion or insertion of nucleotides; the nucleotide similarities of S1, S2, 3a, 3b and E genes among the 27 isolates were 75.9%–100.0%, 85%–100.0%, 64.0%–100.0%, 60.4%–100.0% and 83.1%–100.0%, respectively. According to phylogenetic analysis of S1 gene, the 27 isolates were divided into five genotypes, Mass, Korean-I (K-I), QX-like, KM91-like and New cluster 1. The phylogenetic trees based on the S2, 3a, 3b, E genes and S1-S2-3a-3b-E (S1-E) region nucleotide sequences did not closely follow the clustering based on the S1 sequence. The New cluster 1 prevalent during 2009 and 2010 was not found in 2011 but QX-like viruses became prevalent in 2011. The recombination analysis revealed two new S gene recombinants, 11036 and 11052 which might have been derived from recombinations between the New cluster 1 and QX-like viruses and between the K-I and H120 (vaccine) viruses, respectively. In conclusion, multiple IBV genotypes have co-circulated; QX-like viruses have recurred and new recombinants have emerged in Korea. This has enriched molecular epidemiology information of IBV and is useful for the control of IB in Korea. Viruses 2013-01-31 5 2 Article 10.3390/v5020550 550 567 1999-4915 2013-01-31 doi: 10.3390/v5020550 Mei-Lan Mo Seung-Min Hong Hyuk-Joon Kwon Il-Hwan Kim Chang-Seon Song Jae-Hong Kim http://www.mdpi.com/1999-4915/5/2/528 The Arenaviridae is a diverse and growing family of viruses that includes several agents responsible for important human diseases. Despite the importance of this family for public health, particularly in Africa and South America, much of its biology remains poorly understood. However, in recent years significant progress has been made in this regard, particularly relating to the formation and release of new enveloped virions, which is an essential step in the viral lifecycle. While this process is mediated chiefly by the viral matrix protein Z, recent evidence suggests that for some viruses the nucleoprotein (NP) is also required to enhance the budding process. Here we highlight and compare the distinct budding mechanisms of different arenaviruses, concentrating on the role of the matrix protein Z, its known late domain sequences, and the involvement of cellular endosomal sorting complex required for transport (ESCRT) pathway components. Finally we address the recently described roles for the nucleoprotein NP in budding and ribonucleoprotein complex (RNP) incorporation, as well as discussing possible mechanisms related to its involvement. Viruses 2013-01-31 5 2 Review 10.3390/v5020528 528 549 1999-4915 2013-01-31 doi: 10.3390/v5020528 Svenja Wolff Hideki Ebihara Allison Groseth http://www.mdpi.com/1999-4915/5/2/470 Virus infections are a major global public health concern, and only via substantial knowledge of virus pathogenesis and antiviral immune responses can we develop and improve medical treatments, and preventive and therapeutic vaccines. Innate immunity and the shaping of efficient early immune responses are essential for control of viral infections. In order to trigger an efficient antiviral defense, the host senses the invading microbe via pattern recognition receptors (PRRs), recognizing distinct conserved pathogen-associated molecular patterns (PAMPs). The innate sensing of the invading virus results in intracellular signal transduction and subsequent production of interferons (IFNs) and proinflammatory cytokines. Cytokines, including IFNs and chemokines, are vital molecules of antiviral defense regulating cell activation, differentiation of cells, and, not least, exerting direct antiviral effects. Cytokines shape and modulate the immune response and IFNs are principle antiviral mediators initiating antiviral response through induction of antiviral proteins. In the present review, I describe and discuss the current knowledge on early virus–host interactions, focusing on early recognition of virus infection and the resulting expression of type I and type III IFNs, proinflammatory cytokines, and intracellular antiviral mediators. In addition, the review elucidates how targeted stimulation of innate sensors, such as toll-like receptors (TLRs) and intracellular RNA and DNA sensors, may be used therapeutically. Moreover, I present and discuss data showing how current antimicrobial therapies, including antibiotics and antiviral medication, may interfere with, or improve, immune response. Viruses 2013-01-31 5 2 Review 10.3390/v5020470 470 527 1999-4915 2013-01-31 doi: 10.3390/v5020470 Jesper Melchjorsen http://www.mdpi.com/1999-4915/5/2/439 Hepatitis C virus (HCV) is the etiological agent accounting for chronic liver disease in approximately 2–3% of the population worldwide. HCV infection often leads to liver fibrosis and cirrhosis, various metabolic alterations including steatosis, insulin and interferon resistance or iron overload, and development of hepatocellular carcinoma or non-Hodgkin lymphoma. Multiple molecular mechanisms that trigger the emergence and development of each of these pathogenic processes have been identified so far. One of these involves marked induction of a reactive oxygen species (ROS) in infected cells leading to oxidative stress. To date, markers of oxidative stress were observed both in chronic hepatitis C patients and in various in vitro systems, including replicons or stable cell lines expressing viral proteins. The search for ROS sources in HCV-infected cells revealed several mechanisms of ROS production and thus a number of cellular proteins have become targets for future studies. Furthermore, during last several years it has been shown that HCV modifies antioxidant defense mechanisms. The aim of this review is to summarize the present state of art in the field and to try to predict directions for future studies. Viruses 2013-01-28 5 2 Review 10.3390/v5020439 439 469 1999-4915 2013-01-28 doi: 10.3390/v5020439 Alexander Ivanov Birke Bartosch Olga Smirnova Maria Isaguliants Sergey Kochetkov http://www.mdpi.com/1999-4915/5/2/423 Autoimmune hepatitis (AIH) and primary biliary cirrhosis (PBC) are poorly understood autoimmune liver diseases. Immunosuppression is used to treat AIH and ursodeoxycholic acid is used to slow the progression of PBC. Nevertheless, a proportion of patients with both disorders progress to liver failure. Following liver transplantation, up to a third of patients with PBC experience recurrent disease. Moreover a syndrome referred to as “de novo AIH” occurs in a proportion of patients regardless of maintenance immunosuppression, who have been transplanted for disorders unrelated to AIH. Of note, the use of cyclosporine A appears to protect against the development of recurrent PBC and de novo AIH even though it is a less potent immunosuppressive compared to tacrolimus. The reason why cyclosporine A is protective has not been determined. However, a virus resembling mouse mammary tumor virus (MMTV) has been characterized in patients with PBC and AIH. Accordingly, we hypothesized that the protective effect of cyclosporine A in liver transplant recipients may be mediated by the antiviral activity of this cyclophilin inhibitor. Treatment of the MMTV producing MM5MT cells with different antivirals and immunosuppressive agents showed that both cyclosporine A and the analogue NIM811 inhibited MMTV production from the producer cells. Herein, we discuss the evidence supporting the role of MMTV-like human betaretrovirus in the development of PBC and de novo AIH and speculate on the possibility that the agent may be associated with disease following transplantation. We also review the mechanisms of how both cyclosporine A and NIM811 may inhibit betaretrovirus production in vitro. Viruses 2013-01-24 5 2 Review 10.3390/v5020423 423 438 1999-4915 2013-01-24 doi: 10.3390/v5020423 Shawn Wasilenko Aldo Montano-Loza Andrew Mason http://www.mdpi.com/1999-4915/5/1/406 Pneumovirus infections cause a wide spectrum of respiratory disease in humans and animals. The airway epithelium is the major site of pneumovirus replication. Apoptosis or regulated cell death, may contribute to the host anti-viral response by limiting viral replication. However, apoptosis of lung epithelial cells may also exacerbate lung injury, depending on the extent, the timing and specific location in the lungs. Differential apoptotic responses of epithelial cells versus innate immune cells (e.g., neutrophils, macrophages) during pneumovirus infection can further contribute to the complex and delicate balance between host defense and disease pathogenesis. The purpose of this manuscript is to give an overview of the role of apoptosis in pneumovirus infection. We will examine clinical and experimental data concerning the various pro-apoptotic stimuli and the roles of apoptotic epithelial and innate immune cells during pneumovirus disease. Finally, we will discuss potential therapeutic interventions targeting apoptosis in the lungs. Viruses 2013-01-23 5 1 Review 10.3390/v5010406 406 422 1999-4915 2013-01-23 doi: 10.3390/v5010406 Elske van den Berg Job van Woensel Reinout Bem http://www.mdpi.com/1999-4915/5/1/374 Prion diseases or transmissible spongiform encephalopathies (TSEs) are fatal neurodegenerative disorders in mammals that are caused by unconventional agents predominantly composed of aggregated misfolded prion protein (PrP). Prions self-propagate by recruitment of host-encoded PrP into highly ordered b-sheet rich aggregates. Prion strains differ in their clinical, pathological and biochemical characteristics and are likely to be the consequence of distinct abnormal prion protein conformers that stably replicate their alternate states in the host cell. Understanding prion cell biology is fundamental for identifying potential drug targets for disease intervention. The development of permissive cell culture models has greatly enhanced our knowledge on entry, propagation and dissemination of TSE agents. However, despite extensive research, the precise mechanism of prion infection and potential strain effects remain enigmatic. This review summarizes our current knowledge of the cell biology and propagation of prions derived from cell culture experiments. We discuss recent findings on the trafficking of cellular and pathologic PrP, the potential sites of abnormal prion protein synthesis and potential co-factors involved in prion entry and propagation. Viruses 2013-01-22 5 1 Review 10.3390/v5010374 374 405 1999-4915 2013-01-22 doi: 10.3390/v5010374 Andrea Grassmann Hanna Wolf Julia Hofmann James Graham Ina Vorberg http://www.mdpi.com/1999-4915/5/1/352 Influenza A virus (IAV) has caused seasonal influenza epidemics and influenza pandemics, which resulted in serious threat to public health and socioeconomic impacts. Until now, only 5 drugs belong to two categories are used for prophylaxis and treatment of IAV infection. Hemagglutinin (HA), the envelope glycoprotein of IAV, plays a critical role in viral binding, fusion and entry. Therefore, HA is an attractive target for developing anti‑IAV drugs to block the entry step of IAV infection. Here we reviewed the recent progress in the study of conformational changes of HA during viral fusion process and the development of HA-based IAV entry inhibitors, which may provide a new choice for controlling future influenza pandemics. Viruses 2013-01-22 5 1 Review 10.3390/v5010352 352 373 1999-4915 2013-01-22 doi: 10.3390/v5010352 Jie Yang Minmin Li Xintian Shen Shuwen Liu http://www.mdpi.com/1999-4915/5/1/340 Viral hemorrhagic fevers (VHFs) caused by arenaviruses are acute diseases characterized by fever, headache, general malaise, impaired cellular immunity, eventual neurologic involvement, and hemostatic alterations that may ultimately lead to shock and death. The causes of the bleeding are still poorly understood. However, it is generally accepted that these causes are associated to some degree with impaired hemostasis, endothelial cell dysfunction and low platelet counts or function. In this article, we present the current knowledge about the hematological alterations present in VHF induced by arenaviruses, including new aspects on the underlying pathogenic mechanisms. Viruses 2013-01-21 5 1 Review 10.3390/v5010340 340 351 1999-4915 2013-01-21 doi: 10.3390/v5010340 Mirta Schattner Leonardo Rivadeneyra Roberto Pozner Ricardo Gómez http://www.mdpi.com/1999-4915/5/1/321 While vectored vaccines, based on hyperattenuated viruses, may lead to new treatment options against infectious diseases and certain cancers, they are also complex products and sometimes difficult to provide in sufficient amount and purity. To facilitate vaccine programs utilizing host-restricted poxviruses, we established avian suspension cell lines (CR and CR.pIX) and developed a robust, chemically defined, culturing process for production of this class of vectors. For one prominent member, modified vaccinia Ankara (MVA), we now describe a new strain that appears to replicate to greater yields of infectious units, especially in the cell-free supernatant of cultures in chemically defined media. The new strain was obtained by repeated passaging in CR suspension cultures and, consistent with reports on the exceptional genetic stability of MVA, sequencing of 135 kb of the viral genomic DNA revealed that only three structural proteins (A3L, A9L and A34R) each carry a single amino acid exchange (H639Y, K75E and D86Y, respectively). Host restriction in a plaque-purified isolate of the new genotype appears to be maintained in cell culture. Processing towards an injectable vaccine preparation may be simplified with this strain as a complete lysate, containing the main burden of host cell contaminants, may not be required anymore to obtain adequate yields. Viruses 2013-01-21 5 1 Article 10.3390/v5010321 321 339 1999-4915 2013-01-21 doi: 10.3390/v5010321 Ingo Jordan Deborah Horn Katrin John Volker Sandig http://www.mdpi.com/1999-4915/5/1/295 Respiratory syncytial virus (RSV) is the leading cause of infant bronchiolitis. The closely related pneumonia virus of mice (PVM) causes a similar immune-mediated disease in mice, which allows an analysis of host factors that lead to severe illness. This project was designed to compare the immune responses to lethal and sublethal doses of PVM strain 15 in Balb/c and C57Bl/6 mice. Balb/c mice responded to PVM infection with an earlier and stronger innate response that failed to control viral replication. Production of inflammatory cyto- and chemokines, as well as infiltration of neutrophils and IFN-γ secreting natural killer cells into the lungs, was more predominant in Balb/c mice. In contrast, C57Bl/6 mice were capable of suppressing both viral replication and innate inflammatory responses. After a sublethal infection, PVM-induced IFN-γ production by splenocytes was stronger early during infection and weaker at late time points in C57Bl/6 mice when compared to Balb/c mice. Furthermore, although the IgG levels were similar and the mucosal IgA titres lower, the virus neutralizing antibody titres were higher in C57Bl/6 mice than in Balb/c mice. Overall, the difference in susceptibility of these two strains appeared to be related not to an inherent T helper bias, but to the capacity of the C57Bl/6 mice to control both viral replication and the immune response elicited by PVM. Viruses 2013-01-21 5 1 Article 10.3390/v5010295 295 320 1999-4915 2013-01-21 doi: 10.3390/v5010295 Ellen Watkiss Pratima Shrivastava Natasa Arsic Susantha Gomis Sylvia van Drunen Littel-van den Hurk http://www.mdpi.com/1999-4915/5/1/279 In previous studies, differences in the amount of genomic and subgenomic RNA produced by coronaviruses with mutations in the programmed ribosomal frameshift signal of ORF1a/b were observed. It was not clear if these differences were due to changes in genomic sequence, the protein sequence or the frequency of frameshifting. Here, viruses with synonymous codon changes are shown to produce different ratios of genomic and subgenomic RNA. These findings demonstrate that the protein sequence is not the primary cause of altered genomic and subgenomic RNA production. The synonymous codon changes affect both the structure of the frameshift signal and frameshifting efficiency. Small differences in frameshifting efficiency result in dramatic differences in genomic RNA production and TCID50 suggesting that the frameshifting frequency must stay above a certain threshold for optimal virus production. The data suggest that either the RNA sequence or the ratio of viral proteins resulting from different levels of frameshifting affects viral replication. Viruses 2013-01-18 5 1 Article 10.3390/v5010279 279 294 1999-4915 2013-01-18 doi: 10.3390/v5010279 Ewan Plant Amy Sims Ralph Baric Jonathan Dinman Deborah Taylor http://www.mdpi.com/1999-4915/5/1/241 Arenavirus particles are enveloped and contain two single-strand RNA genomic segments with ambisense coding. Genetic plasticity of the arenaviruses comes from transcription errors, segment reassortment, and permissive genomic packaging, and results in their remarkable ability, as a group, to infect a wide variety of hosts. In this review, we discuss some in vitro studies of virus genetic and phenotypic variation after exposure to selective pressures such as high viral dose, mutagens and antivirals. Additionally, we discuss the variation in vivo of selected isolates of Old World arenaviruses, particularly after infection of different animal species. We also discuss the recent emergence of new arenaviruses in the context of our observations of sequence variations that appear to be host-specific. Viruses 2013-01-17 5 1 Review 10.3390/v5010241 241 278 1999-4915 2013-01-17 doi: 10.3390/v5010241 Juan Zapata Maria Salvato http://www.mdpi.com/1999-4915/5/1/226 Epstein-Barr Nuclear Antigen 1 (EBNA1) is a multifunctional protein encoded by EBV. EBNA1’s role in maintaining EBV in latently proliferating cells, by mediating EBV genome synthesis and nonrandom partitioning to daughter cells, as well as regulating viral gene transcription, is well characterized. Less understood are the roles of EBNA1 in affecting the host cell to provide selective advantages to those cells that harbor EBV. In this review we will focus on the interactions between EBNA1 and the host cell that may provide EBV-infected cells selective advantages beyond the maintenance of EBV. Viruses 2013-01-16 5 1 Review 10.3390/v5010226 226 240 1999-4915 2013-01-16 doi: 10.3390/v5010226 Danielle Smith Bill Sugden http://www.mdpi.com/1999-4915/5/1/211 Human respiratory syncytial virus (RSV) is the main viral cause of respiratory tract infection in infants as well as some elderly and high-risk adults with chronic pulmonary disease and the severely immunocompromised. So far, no specific anti-RSV therapeutics or effective anti-RSV vaccines have been reported. Only one humanized monoclonal antibody, Palivizumab, has been approved for use in high-risk infants to prevent RSV infection. Ribavirin is the only drug licensed for therapy of RSV infection, but its clinical use is limited by its nonspecific anti-RSV activity, toxic effect, and relatively high cost. Therefore, development of novel effective anti-RSV therapeutics is urgently needed. The RSV envelope glycoprotein F plays an important role in RSV fusion with, and entry into, the host cell and, consequently, serves as an attractive target for developing RSV entry inhibitors. This article reviews advances made in studies of the structure and function of the F protein and the development of RSV entry inhibitors targeting it. Viruses 2013-01-16 5 1 Review 10.3390/v5010211 211 225 1999-4915 2013-01-16 doi: 10.3390/v5010211 Zhiwu Sun Yanbin Pan Shibo Jiang Lu Lu http://www.mdpi.com/1999-4915/5/1/192 Human metapneumovirus (HMPV) is a leading cause of respiratory infection that causes upper airway and severe lower respiratory tract infections. HMPV infection is initiated by viral surface glycoproteins that attach to cellular receptors and mediate virus membrane fusion with cellular membranes. Most paramyxoviruses use two viral glycoproteins to facilitate virus entry—an attachment protein and a fusion (F) protein. However, membrane fusion for the human paramyxoviruses in the Pneumovirus subfamily, HMPV and respiratory syncytial virus (hRSV), is unique in that the F protein drives fusion in the absence of a separate viral attachment protein. Thus, pneumovirus F proteins can perform the necessary functions for virus entry, i.e., attachment and fusion. In this review, we discuss recent advances in the understanding of how HMPV F mediates both attachment and fusion. We review the requirements for HMPV viral surface glycoproteins during entry and infection, and review the identification of cellular receptors for HMPV F. We also review our current understanding of how HMPV F mediates fusion, concentrating on structural regions of the protein that appear to be critical for membrane fusion activity. Finally, we illuminate key unanswered questions and suggest how further studies can elucidate how this clinically important paramyxovirus fusion protein may have evolved to initiate infection by a unique mechanism. Viruses 2013-01-16 5 1 Review 10.3390/v5010192 192 210 1999-4915 2013-01-16 doi: 10.3390/v5010192 Reagan Cox John Williams http://www.mdpi.com/1999-4915/5/1/182 Cyclophilin A (CypA) is the main member of the immunophilin superfamily that has peptidyl-prolyl cis-trans isomerase activity. CypA participates in protein folding, cell signaling, inflammation and tumorigenesis. Further, CypA plays critical roles in the replication of several viruses. Upon influenza virus infection, CypA inhibits viral replication by interacting with the M1 protein. In addition, CypA is incorporated into the influenza virus virions. Finally, Cyclosporin A (CsA), the main inhibitor of CypA, inhibits influenza virus replication through CypA-dependent and -independent pathways. This review briefly summarizes recent advances in understanding the roles of CypA during influenza virus infection. Viruses 2013-01-15 5 1 Review 10.3390/v5010182 182 191 1999-4915 2013-01-15 doi: 10.3390/v5010182 Xiaoling Liu Zhendong Zhao Wenjun Liu http://www.mdpi.com/1999-4915/5/1/162 Viral codon usage bias may be the product of a number of synergistic or antagonistic factors, including genomic nucleotide composition, translational selection, genomic architecture, and mutational or repair biases. Most studies of viral codon bias evaluate only the relative importance of genomic base composition and translational selection, ignoring other possible factors. We analyzed the codon preferences of ssRNA (luteoviruses and potyviruses) and ssDNA (geminiviruses) plant viruses that infect translationally distinct monocot and dicot hosts. We found that neither genomic base composition nor translational selection satisfactorily explains their codon usage biases. Furthermore, we observed a strong relationship between the codon preferences of viruses in the same family or genus, regardless of host or genomic nucleotide content. Our results suggest that analyzing codon bias as either due to base composition or translational selection is a false dichotomy that obscures the role of other factors. Constraints such as genomic architecture and secondary structure can and do influence codon usage in plant viruses, and likely in viruses of other hosts. Viruses 2013-01-15 5 1 Article 10.3390/v5010162 162 181 1999-4915 2013-01-15 doi: 10.3390/v5010162 Daniel Cardinale Kate DeRosa Siobain Duffy http://www.mdpi.com/1999-4915/5/1/150 In this study, three-day old mature biofilms of Escherichia coli were exposed once to either a temperate Shiga-toxin encoding phage (H-19B) or an obligatory lytic phage (T7), after which further dynamics in the biofilm were monitored. As such, it was found that a single dose of H-19B could rapidly lead to a near complete lysogenization of the biofilm, with a subsequent continuous release of infectious H-19B particles. On the other hand, a single dose of T7 rapidly led to resistance development in the biofilm population. Together, our data indicates a profound impact of phages on the dynamics within structured bacterial populations. Viruses 2013-01-11 5 1 Communication 10.3390/v5010150 150 161 1999-4915 2013-01-11 doi: 10.3390/v5010150 Pieter Moons David Faster Abram Aertsen http://www.mdpi.com/1999-4915/5/1/127 The hydrophobic pocket in the HIV-1 gp41 N-terminal heptad repeat (NHR) domain plays an important role in viral fusion and entry into the host cell, and serves as an attractive target for development of HIV-1 fusion/entry inhibitors. The peptide anti-HIV drug targeting gp41 NHR, T-20 (generic name: enfuvirtide; brand name: Fuzeon), was approved by the U.S. FDA in 2003 as the first HIV fusion/entry inhibitor for treatment of HIV/AIDS patients who fail to respond to the current antiretroviral drugs. However, because T20 lacks the pocket-binding domain (PBD), it exhibits low anti-HIV-1 activity and short half-life. Therefore, several next-generation HIV fusion inhibitory peptides with PBD have been developed. They possess longer half-life and more potent antiviral activity against a broad spectrum of HIV-1 strains, including the T-20-resistant variants. Nonetheless, the clinical application of these peptides is still limited by the lack of oral availability and the high cost of production. Thus, development of small molecule compounds targeting the gp41 pocket with oral availability has been promoted. This review describes the main approaches for identification of HIV fusion/entry inhibitors targeting the gp41 pocket and summarizes the latest progress in developing these inhibitors as a new class of anti-HIV drugs. Viruses 2013-01-11 5 1 Review 10.3390/v5010127 127 149 1999-4915 2013-01-11 doi: 10.3390/v5010127 Fei Yu Lu Lu Lanying Du Xiaojie Zhu Asim Debnath Shibo Jiang http://www.mdpi.com/1999-4915/5/1/111 In a λimm434 lysogen, two proteins are expressed from the integrated prophage. Both are encoded by the same mRNA whose transcription initiates at the PRM promoter. One protein is the 434 repressor, needed for the establishment and maintenance of lysogeny. The other is Hex which is translated from an open reading frame that apparently partially overlaps the 434 repressor coding region. In the wild type host, disruption of the gene encoding Hex destabilizes λimm434 lysogens. However, the hex mutation has no effect on lysogen stability in a recA− host. These observations suggest that Hex functions by modulating the ability of RecA to stimulate 434 repressor autocleavage. We tested this hypothesis by identifying and purifying Hex to determine if this protein inhibited RecA‑stimulated autocleavage of 434 repressor in vitro. Our results show that in vitro a fragment of Hex prevents RecA-stimulated autocleavage of 434 repressor, as well as the repressors of the closely related phage P22. Surprisingly, Hex does not prevent RecA‑stimulated autocleavage of phage lambda repressor, nor the E. coli LexA repressor. Viruses 2013-01-09 5 1 Article 10.3390/v5010111 111 126 1999-4915 2013-01-09 doi: 10.3390/v5010111 Paul Shkilnyj Michael Colon Gerald Koudelka http://www.mdpi.com/1999-4915/5/1/87 Human metapneumovirus (HMPV) is a relative newly described virus. It was first isolated in 2001 and currently appears to be one of the most significant and common human viral infections. Retrospective serologic studies demonstrated the presence of HMPV antibodies in humans more than 50 years earlier. Although the virus was primarily known as causative agent of respiratory tract infections in children, HMPV is an important cause of respiratory infections in adults as well. Almost all children are infected by HMPV below the age of five; the repeated infections throughout life indicate transient immunity. HMPV infections usually are mild and self-limiting, but in the frail elderly and the immunocompromised patients, the clinical course can be complicated. Since culturing the virus is relatively difficult, diagnosis is mostly based on a nucleic acid amplification test, such as reverse transcriptase polymerase chain reaction. To date, no vaccine is available and treatment is supportive. However, ongoing research shows encouraging results. The aim of this paper is to review the current literature concerning HMPV infections in adults, and discuss recent development in treatment and vaccination. Viruses 2013-01-08 5 1 Review 10.3390/v5010087 87 110 1999-4915 2013-01-08 doi: 10.3390/v5010087 Lenneke Haas Steven Thijsen Leontine van Elden Karen Heemstra http://www.mdpi.com/1999-4915/5/1/54 Viral polymerases are important targets in drug discovery and development efforts. Most antiviral compounds that are currently approved for treatment of infection with members of the herpesviridae family were shown to inhibit the viral DNA polymerase. However, biochemical studies that shed light on mechanisms of drug action and resistance are hampered primarily due to technical problems associated with enzyme expression and purification. In contrast, the orthologous bacteriophage RB69 polymerase gp43 has been crystallized in various forms and therefore serves as a model system that provides a better understanding of structure–function relationships of polymerases that belong the type B family. This review aims to discuss strengths, limitations, and opportunities of the phage surrogate with emphasis placed on its utility in the discovery and development of anti-herpetic drugs. Viruses 2013-01-08 5 1 Review 10.3390/v5010054 54 86 1999-4915 2013-01-08 doi: 10.3390/v5010054 Nicholas Bennett Matthias Götte http://www.mdpi.com/1999-4915/5/1/15 Pseudomonas aeruginosa is a frequent participant in wound infections. Emergence of multiple antibiotic resistant strains has created significant problems in the treatment of infected wounds. Phage therapy (PT) has been proposed as a possible alternative approach. Infected wounds are the perfect place for PT applications, since the basic condition for PT is ensured; namely, the direct contact of bacteria and their viruses. Plenty of virulent (“lytic”) and temperate (“lysogenic”) bacteriophages are known in P. aeruginosa. However, the number of virulent phage species acceptable for PT and their mutability are limited. Besides, there are different deviations in the behavior of virulent (and temperate) phages from their expected canonical models of development. We consider some examples of non-canonical phage-bacterium interactions and the possibility of their use in PT. In addition, some optimal approaches to the development of phage therapy will be discussed from the point of view of a biologist, considering the danger of phage-assisted horizontal gene transfer (HGT), and from the point of view of a surgeon who has accepted the Hippocrates Oath to cure patients by all possible means. It is also time now to discuss the possible approaches in international cooperation for the development of PT. We think it would be advantageous to make phage therapy a kind of personalized medicine. Viruses 2012-12-21 5 1 Review 10.3390/v5010015 15 53 1999-4915 2012-12-21 doi: 10.3390/v5010015 Victor Krylov Olga Shaburova Sergey Krylov Elena Pleteneva http://www.mdpi.com/1999-4915/5/1/1 The expression of intrinsic antiviral factors by myeloid cells is a recently recognized mechanism of restricting lentiviral replication. Viruses that enter these cells must develop strategies to evade cellular antiviral factors to establish a productive infection. By studying the cellular targets of virally encoded proteins that are necessary to infect myeloid cells, a better understanding of cellular intrinsic antiviral strategies has now been achieved. Recent findings have provided insight into how the lentiviral accessory proteins, Vpx, Vpr and Vif counteract antiviral factors found in myeloid cells including SAMHD1, APOBEC3G, APOBEC3A, UNG2 and uracil. Here we review our current understanding of the molecular basis of how cellular antiviral factors function and the viral countermeasures that antagonize them to promote viral transmission and spread. Viruses 2012-12-21 5 1 Review 10.3390/v5010001 1 14 1999-4915 2012-12-21 doi: 10.3390/v5010001 Mike Mashiba Kathleen Collins http://www.mdpi.com/1999-4915/4/12/3952 It has come to my attention that my article "Latest Insights on Adenovirus Structure and Assembly" (Viruses 2012, 4, 847-877) [1] contains an inaccurate statement. On page 864, the caption for Figure 7 reads: "There are four potential cleavage sites in pTP but they have not been experimentally verified". However, three of these sites have been experimentally confirmed in vitro using recombinant AVP and pTP, as described in Webster A, Leith I.R., Hay R.T.: Activation of adenovirus-coded protease and processing of preterminal protein. J. Virol. 1994, 68, 7292-7300 [2]. Viruses 2012-12-19 4 12 Correction 10.3390/v4123952 3952 3952 1999-4915 2012-12-19 doi: 10.3390/v4123952 Carmen Martín http://www.mdpi.com/1999-4915/4/12/3932 We describe herein, limitations on research at biosafety level 4 (BSL-4) containment laboratories, with regard to biosecurity regulations, safety considerations, research space limitations, and physical constraints in executing experimental procedures. These limitations can severely impact the number of collaborations and size of research projects investigating microbial pathogens of biodefense concern. Acquisition, use, storage, and transfer of biological select agents and toxins (BSAT) are highly regulated due to their potential to pose a severe threat to public health and safety. All federal, state, city, and local regulations must be followed to obtain and maintain registration for the institution to conduct research involving BSAT. These include initial screening and continuous monitoring of personnel, controlled access to containment laboratories, accurate and current BSAT inventory records. Safety considerations are paramount in BSL-4 containment laboratories while considering the types of research tools, workflow and time required for conducting both in vivo and in vitro experiments in limited space. Required use of a positive-pressure encapsulating suit imposes tremendous physical limitations on the researcher. Successful mitigation of these constraints requires additional time, effort, good communication, and creative solutions. Test and evaluation of novel vaccines and therapeutics conducted under good laboratory practice (GLP) conditions for FDA approval are prioritized and frequently share the same physical space with important ongoing basic research studies. The possibilities and limitations of biomedical research involving microbial pathogens of biodefense concern in BSL-4 containment laboratories are explored in this review. Viruses 2012-12-19 4 12 Review 10.3390/v4123932 3932 3951 1999-4915 2012-12-19 doi: 10.3390/v4123932 Amy Shurtleff Nicole Garza Matthew Lackemeyer Ricardo Carrion Anthony Griffiths Jean Patterson Samuel Edwin Sina Bavari http://www.mdpi.com/1999-4915/4/12/3912 Liver diseases, such as hepatitis C virus (HCV), are “broken spirit” diseases. The prevalence of HCV infection for American Indian/Alaskan Native (AI/AN) in the United States and Canadian Aboriginals varies; nonetheless, incidence rates of newly diagnosed HCV infection are typically higher relative to non-indigenous people. For AI/AN and Aboriginal peoples risk factors for the diagnosis of HCV can reflect that of the general population: predominately male, a history of injection drug use, in midlife years, with a connection with urban centers. However, the face of the indigenous HCV infected individual is becoming increasingly female and younger compared to non-indigenous counterparts. Epidemiology studies indicate that more effective clearance of acute HCV infection can occur for select Aboriginal populations, a phenomenon which may be linked to unique immune characteristics. For individuals progressing to chronic HCV infection treatment outcomes are comparable to other racial cohorts. Disease progression, however, is propelled by elevated rates of co-morbidities including type 2 diabetes and alcohol use, along with human immunodeficiency virus (HIV) co-infection relative to non-indigenous patients. Historical and personal trauma has a major role in the participation of high risk behaviors and associated diseases. Although emerging treatments provide hope, combating HCV related morbidity and mortality will require interventions that address the etiology of broken spirit diseases. Viruses 2012-12-19 4 12 Review 10.3390/v4123912 3912 3931 1999-4915 2012-12-19 doi: 10.3390/v4123912 Julia Rempel Julia Uhanova http://www.mdpi.com/1999-4915/4/12/3859 The human immunodeficiency virus (HIV) enters cells through a series of molecular interactions between the HIV envelope protein and cellular receptors, thus providing many opportunities to block infection. Entry inhibitors are currently being used in the clinic, and many more are under development. Unfortunately, as is the case for other classes of antiretroviral drugs that target later steps in the viral life cycle, HIV can become resistant to entry inhibitors. In contrast to inhibitors that block viral enzymes in intracellular compartments, entry inhibitors interfere with the function of the highly variable envelope glycoprotein as it continuously adapts to changing immune pressure and available target cells in the extracellular environment. Consequently, pathways and mechanisms of resistance for entry inhibitors are varied and often involve mutations across the envelope gene. This review provides a broad overview of entry inhibitor resistance mechanisms that inform our understanding of HIV entry and the design of new inhibitors and vaccines. Viruses 2012-12-19 4 12 Review 10.3390/v4123859 3859 3911 1999-4915 2012-12-19 doi: 10.3390/v4123859 Christopher De Feo Carol Weiss http://www.mdpi.com/1999-4915/4/12/3851 Human Papillomavirus (HPV) infection is the most common sexually transmitted infection (STI) worldwide. Incidence rates of HPV infection among human immunodeficiency virus (HIV)-infected individuals are well documented and are several-fold higher than among HIV-uninfected individuals. Few studies have demonstrated an increased risk for acquiring HIV infection in those with HPV infection, and this risk seems to be higher when HPV strains are of high-risk oncogenic potential. The estimated prevalence of high-risk oncogenic HPV infection is highest in men who have sex with men (MSM), a particularly vulnerable group with high prevalence rates of HIV infection and other STIs. In this paper, we provide a comprehensive review of the available literature on the role of HPV infection in HIV acquisition. Our review includes data from cross-sectional and longitudinal studies. Viruses 2012-12-18 4 12 Review 10.3390/v4123851 3851 3858 1999-4915 2012-12-18 doi: 10.3390/v4123851 Brandon Brown Mariam Davtyan Jerome Galea Erica Chow Segundo Leon Jeffrey Klausner http://www.mdpi.com/1999-4915/4/12/3831 The ability of the host to trigger apoptosis in infected cells is perhaps the most powerful tool by which viruses can be cleared from the host organism. To avoid elimination by this mechanism, human papillomaviruses (HPV) have developed several mechanisms that enable the cells they infect to elude both extrinsic and intrinsic apoptosis. In this manuscript, we review the current literature regarding how HPV-infected cells avoid apoptosis and the molecular mechanisms involved in these events. In particular, we will discuss the modifications in intrinsic and extrinsic apoptotic pathways caused by proteins encoded by HPV early genes. Many of the current efforts regarding anti-cancer drug development are focused on directing tumor cells to undergo apoptosis. However, the ability of HPV-infected cells to resist apoptotic signals renders such therapies ineffective. Possible mechanisms for overcoming the resistance of HPV-infected tumor cells to anticancer drugs will be discussed. Viruses 2012-12-18 4 12 Review 10.3390/v4123831 3831 3850 1999-4915 2012-12-18 doi: 10.3390/v4123831 Chung-Hsiang Yuan Maria Filippova Penelope Duerksen-Hughes http://www.mdpi.com/1999-4915/4/12/3812 West Nile virus (WNV) continues to cause outbreaks of severe neuroinvasive disease in humans and other vertebrate animals in the United States, Europe, and other regions of the world. This review discusses our understanding of the interactions between virus and host that occur in the central nervous system (CNS), the outcome of which can be protection, viral pathogenesis, or immunopathogenesis. We will focus on defining the current state of knowledge of WNV entry, tropism, and host immune response in the CNS, all of which affect the balance between injury and successful clearance. Viruses 2012-12-17 4 12 Review 10.3390/v4123812 3812 3830 1999-4915 2012-12-17 doi: 10.3390/v4123812 Hyelim Cho Michael Diamond http://www.mdpi.com/1999-4915/4/12/3804 Yardlong bean (Vigna unguiculata subsp. sesquipedalis) plants with virus-like systemic mottling and leaf distortion were observed in both experimental and commercial fields in Aragua State, Venezuela. Symptomatic leaves were shown to contain carlavirus-like particles. RT-PCR analysis with carlavirus-specific primers was positive in all tested samples. Nucleotide sequences of the obtained amplicons showed 84%–74% similarity to corresponding sequences of Cowpea mild mottle virus (CPMMV) isolates deposited in the GenBank database. This is the first report of CPMMV in Venezuela and is thought to be the first report of CPMMV infecting yardlong bean. Viruses 2012-12-14 4 12 Article 10.3390/v4123804 3804 3811 1999-4915 2012-12-14 doi: 10.3390/v4123804 Miriam Brito Thaly Fernández-Rodríguez Mario Garrido Alexander Mejías Mirtha Romano Edgloris Marys http://www.mdpi.com/1999-4915/4/12/3785 We have previously shown that hMPV G protein (B2 lineage) interacts with cellular glycosaminoglycans (GAGs). In this study we examined subtypes A1, A2 and B1 for this interaction. GAG-dependent infectivity of available hMPV strains was demonstrated using GAG-deficient cells and heparin competition. We expressed the G protein ectodomains from all strains and analysed these by heparin affinity chromatography. In contrast to the B2 lineage, neither the A2 or B1 G proteins bound to heparin. Sequence analysis of these strains indicated that although there was some homology with the B2 heparin-binding domains, there were less positively charged residues, providing a likely explanation for the lack of binding. Although sequence analysis did not demonstrate well defined positively charged domains in G protein of the A1 strain, this protein was able to bind heparin, albeit with a lower affinity than G protein of the B2 strain. These results indicate diversity in GAG interactions between G proteins of different lineages and suggest that the GAG-dependency of all strains may be mediated by interaction with an alternative surface protein, most probably the conserved fusion (F) protein. Analysis of both native and recombinant F protein confirmed that F protein binds heparin, supporting this conclusion. Viruses 2012-12-14 4 12 Article 10.3390/v4123785 3785 3803 1999-4915 2012-12-14 doi: 10.3390/v4123785 Penelope Adamson Sutthiwan Thammawat Gamaliel Muchondo Tania Sadlon David Gordon http://www.mdpi.com/1999-4915/4/12/3754 Historically, mice and guinea pigs have been the rodent models of choice for therapeutic and prophylactic countermeasure testing against Ebola virus disease (EVD). Recently, hamsters have emerged as a novel animal model for the in vivo study of EVD. In this review, we discuss the history of the hamster as a research laboratory animal, as well as current benefits and challenges of this model. Availability of immunological reagents is addressed. Salient features of EVD in hamsters, including relevant pathology and coagulation parameters, are compared directly with the mouse, guinea pig and nonhuman primate models. Viruses 2012-12-14 4 12 Review 10.3390/v4123754 3754 3784 1999-4915 2012-12-14 doi: 10.3390/v4123754 Victoria Wahl-Jensen Laura Bollinger David Safronetz Fabian de Kok-Mercado Dana Scott Hideki Ebihara http://www.mdpi.com/1999-4915/4/12/3731 Respiratory syncytial virus (RSV) is the most common viral cause of childhood acute lower respiratory tract infections. It is estimated that RSV infections result in more than 100,000 deaths annually worldwide. Bovine RSV is a cause of enzootic pneumonia in young dairy calves and summer pneumonia in nursing beef calves. Furthermore, bovine RSV plays a significant role in bovine respiratory disease complex, the most prevalent cause of morbidity and mortality among feedlot cattle. Infection of calves with bovine RSV shares features in common with RSV infection in children, such as an age-dependent susceptibility. In addition, comparable microscopic lesions consisting of bronchiolar neutrophilic infiltrates, epithelial cell necrosis, and syncytial cell formation are observed. Further, our studies have shown an upregulation of pro-inflammatory mediators in RSV-infected calves, including IL-12p40 and CXCL8 (IL-8). This finding is consistent with increased levels of IL-8 observed in children with RSV bronchiolitis. Since rodents lack IL-8, neonatal calves can be useful for studies of IL-8 regulation in response to RSV infection. We have recently found that vitamin D in milk replacer diets can be manipulated to produce calves differing in circulating 25-hydroxyvitamin D3. The results to date indicate that although the vitamin D intracrine pathway is activated during RSV infection, pro-inflammatory mediators frequently inhibited by the vitamin D intacrine pathway in vitro are, in fact, upregulated or unaffected in lungs of infected calves. This review will summarize available data that provide parallels between bovine RSV infection in neonatal calves and human RSV in infants. Viruses 2012-12-13 4 12 Review 10.3390/v4123731 3731 3753 1999-4915 2012-12-13 doi: 10.3390/v4123731 Randy Sacco Jodi McGill Mitchell Palmer John Lippolis Timothy Reinhardt Brian Nonnecke http://www.mdpi.com/1999-4915/4/12/3701 Epidemiological data suggest that the Epstein-Barr virus (EBV) is associated with several autoimmune diseases, such as systemic lupus erythematosus, rheumatoid arthritis and multiple sclerosis. However, it is not clear whether EBV plays a role in the pathogenesis of these diseases, and if so, by which mechanisms the virus may contribute. In this review, we discuss possible viral and immunological mechanisms that might explain associations between EBV and autoimmune diseases and whether these associations represent causes or effects of inflammation and autoimmunity. Viruses 2012-12-13 4 12 Review 10.3390/v4123701 3701 3730 1999-4915 2012-12-13 doi: 10.3390/v4123701 Andreas Lossius Jorunn Johansen Øivind Torkildsen Frode Vartdal Trygve Holmøy http://www.mdpi.com/1999-4915/4/12/3689 In 2007, a novel coronavirus associated with an acute respiratory disease in alpacas (Alpaca Coronavirus, ACoV) was isolated. Full-length genomic sequencing of the ACoV demonstrated the genome to be consistent with other Alphacoronaviruses. A putative additional open-reading frame was identified between the nucleocapsid gene and 3'UTR. The ACoV was genetically most similar to the common human coronavirus (HCoV) 229E with 92.2% nucleotide identity over the entire genome. A comparison of spike gene sequences from ACoV and from HCoV-229E isolates recovered over a span of five decades showed the ACoV to be most similar to viruses isolated in the 1960’s to early 1980’s. The true origin of the ACoV is unknown, however a common ancestor between the ACoV and HCoV-229E appears to have existed prior to the 1960’s, suggesting virus transmission, either as a zoonosis or anthroponosis, has occurred between alpacas and humans. Viruses 2012-12-12 4 12 Article 10.3390/v4123689 3689 3700 1999-4915 2012-12-12 doi: 10.3390/v4123689 Beate Crossley Richard Mock Scott Callison Sharon Hietala http://www.mdpi.com/1999-4915/4/12/3665 In the last 20 years, molecular ecology approaches have proven to be extremely useful to identify and assess factors associated with viral emerging diseases, particularly in economically and socially important tropical crops such as maize (maize streak disease) and cassava (cassava mosaic disease). Molecular ecology approaches were applied in Reunion Island to analyze the epidemic of tomato yellow leaf curl disease, which has been affecting the island since the end of the 1990s. Before the invasive biotype B (currently known as Middle East-Asia Minor 1 cryptic species) of Bemisia tabaci spread across the world, Reunion Island (South West Indian Ocean) only hosted an indigenous biotype of B. tabaci, Ms (currently known as Indian Ocean cryptic species). Wild hybrids between invasive and indigenous species were subsequently characterized over multiple generations. Endosymbiont analysis of the hybrid population indicated that matings were non-random. Similarly, while no indigenous begomoviruses have ever been reported on Reunion Island, the two main strains of one of the most damaging and emerging plant viruses in the world, the Mild and Israel strains of the Tomato yellow leaf curl virus (TYLCV-Mld and TYLCV-IL), were introduced in 1997 and 2004 respectively. While these introductions extensively modified the agricultural landscape of Reunion Island, they also provided an invaluable opportunity to study the ecological and genetic mechanisms involved in biological invasion and competition. Viruses 2012-12-12 4 12 Review 10.3390/v4123665 3665 3688 1999-4915 2012-12-12 doi: 10.3390/v4123665 Frédéric Péréfarres Magali Thierry Nathalie Becker Pierre Lefeuvre Bernard Reynaud Hélène Delatte Jean-Michel Lett http://www.mdpi.com/1999-4915/4/12/3647 Detailed knowledge of the host-virus interactions that accompany filovirus entry into cells is expected to identify determinants of viral virulence and host range, and to yield targets for the development of antiviral therapeutics. While it is generally agreed that filovirus entry into the host cytoplasm requires viral internalization into acidic endosomal compartments and proteolytic cleavage of the envelope glycoprotein by endo/lysosomal cysteine proteases, our understanding of the specific endocytic pathways co-opted by filoviruses remains limited. This review addresses the current knowledge on cellular endocytic pathways implicated in filovirus entry, highlights the consensus as well as controversies, and discusses important remaining questions. Viruses 2012-12-11 4 12 Review 10.3390/v4123647 3647 3664 1999-4915 2012-12-11 doi: 10.3390/v4123647 Suchita Bhattacharyya Nirupama Mulherkar Kartik Chandran http://www.mdpi.com/1999-4915/4/12/3625 Systems biology approaches in virology aim to integrate viral and host biological networks, and thus model the infection process. The growing availability of high-throughput “-omics” techniques and datasets, as well as the ever-increasing sophistication of in silico modeling tools, has resulted in a corresponding rise in the complexity of the analyses that can be performed. The present study seeks to review and organize published evidence regarding virus-host interactions for the arenaviruses, from alterations in the host proteome during infection, to reported protein-protein interactions. In this way, we hope to provide an overview of the interplay between arenaviruses and the host cell, and lay the foundations for complementing current arenavirus research with a systems-level approach. Viruses 2012-12-11 4 12 Review 10.3390/v4123625 3625 3646 1999-4915 2012-12-11 doi: 10.3390/v4123625 Magali Droniou-Bonzom Paula Cannon http://www.mdpi.com/1999-4915/4/12/3606 A novel DNA vaccine vector encoding the Mycobacterium tuberculosis secreted antigen Ag85A fused with the influenza A virus (IAV) HA2 protein epitopes, pEGFP/Ag85A-sHA2 (pAg85A-sHA2), was designed to provide protection against influenza. The antigen encoded by the DNA vaccine vector was efficiently expressed in mammalian cells, as determined by reverse transcription polymerase chain reaction (RT-PCR) and fluorescence analyses. Mice were immunized with the vaccine vector by intramuscular injection before challenge with A/Puerto Rico/8/34 virus (PR8 virus). Sera and the splenocyte culture IFN-γ levels were significantly higher in immunized mice compared with the control mice. The novel vaccine group showed a high neutralization antibody titer in vitro. The novel vaccine vector also reduced the viral loads, increased the survival rates in mice after the PR8 virus challenge and reduced the alveolar inflammatory cell numbers. Sera IL-4 concentrations were significantly increased in mice immunized with the novel vaccine vector on Day 12 after challenge with the PR8 virus. These results demonstrated that short HA2 (sHA2) protein epitopes may provide protection against the PR8 virus and that Ag85A could strengthen the immune response to HA2 epitopes, thus, Ag85A may be developed as a new adjuvant for influenza vaccines. Viruses 2012-12-10 4 12 Article 10.3390/v4123606 3606 3624 1999-4915 2012-12-10 doi: 10.3390/v4123606 Jun Dai Decui Pei Baoning Wang Yu Kuang Laifeng Ren Kang Cao Huan Wang Bin Zuo Jingjing Shao Sha Li Hong Li Mingyuan Li http://www.mdpi.com/1999-4915/4/12/3589 Several global outbreaks of highly pathogenic avian influenza (HPAI) H5N1 virus have increased the urgency of developing effective and safe vaccines against H5N1. Compared with H5N1 inactivated vaccines used widely, H5N1 live-attenuated influenza vaccines (LAIVs) have advantages in vaccine efficacy, dose-saving formula, long-lasting effect, ease of administration and some cross-protective immunity. Furthermore, H5N1 LAIVs induce both humoral and cellular immune responses, especially including improved IgA production at the mucosa. The current trend of H5N1 LAIVs development is toward cold-adapted, temperature-sensitive or replication-defective vaccines, and moreover, H5N1 LAIVs plus mucosal adjuvants are promising candidates. This review provides an update on the advantages and development of H5N1 live-attenuated influenza vaccines. Viruses 2012-12-10 4 12 Review 10.3390/v4123589 3589 3605 1999-4915 2012-12-10 doi: 10.3390/v4123589 Dan Zheng Yinglei Yi Ze Chen http://www.mdpi.com/1999-4915/4/12/3572 Foamy virus (FV) vectors have shown great promise for hematopoietic stem cell (HSC) gene therapy. Their ability to efficiently deliver transgenes to multi-lineage long-term repopulating cells in large animal models suggests they will be effective for several human hematopoietic diseases. Here, we review FV vector studies in large animal models, including the use of FV vectors with the mutant O6-methylguanine-DNA methyltransferase, MGMTP140K to increase the number of genetically modified cells after transplantation. In these studies, FV vectors have mediated efficient gene transfer to polyclonal repopulating cells using short ex vivo transduction protocols designed to minimize the negative effects of ex vivo culture on stem cell engraftment. In this regard, FV vectors appear superior to gammaretroviral vectors, which require longer ex vivo culture to effect efficient transduction. FV vectors have also compared favorably with lentiviral vectors when directly compared in the dog model. FV vectors have corrected leukocyte adhesion deficiency and pyruvate kinase deficiency in the dog large animal model. FV vectors also appear safer than gammaretroviral vectors based on a reduced frequency of integrants near promoters and also near proto-oncogenes in canine repopulating cells. Together, these studies suggest that FV vectors should be highly effective for several human hematopoietic diseases, including those that will require relatively high percentages of gene-modified cells to achieve clinical benefit. Viruses 2012-12-07 4 12 Review 10.3390/v4123572 3572 3588 1999-4915 2012-12-07 doi: 10.3390/v4123572 Grant Trobridge Peter Horn Brian Beard Hans-Peter Kiem http://www.mdpi.com/1999-4915/4/12/3551 Human metapneumovirus (hMPV) is a recently identified RNA virus belonging to the Paramyxoviridae family, which includes several major human and animal pathogens. Epidemiological studies indicate that hMPV is a significant human respiratory pathogen with worldwide distribution. It is associated with respiratory illnesses in children, adults, and immunocompromised patients, ranging from upper respiratory tract infections to severe bronchiolitis and pneumonia. Interferon (IFN) represents a major line of defense against virus infection, and in response, viruses have evolved countermeasures to inhibit IFN production as well as IFN signaling. Although the strategies of IFN evasion are similar, the specific mechanisms by which paramyxoviruses inhibit IFN responses are quite diverse. In this review, we will present an overview of the strategies that hMPV uses to subvert cellular signaling in airway epithelial cells, the major target of infection, as well as in primary immune cells. Viruses 2012-12-07 4 12 Review 10.3390/v4123551 3551 3571 1999-4915 2012-12-07 doi: 10.3390/v4123551 Deepthi Kolli Xiaoyong Bao Antonella Casola http://www.mdpi.com/1999-4915/4/12/3531 The worldwide prevalence of HCV infection is between 1% and 8% in pregnant women and between 0.05% and 5% in children. Yet the pathogenesis of hepatitis C during pregnancy and in the neonatal period remains poorly understood. Mother-to-child transmission (MTCT), a leading cause of pediatric HCV infection, takes place at a rate of <10%. Factors that increase the risk of MTCT include high maternal HCV viral load and coinfection with HIV-1 but, intriguingly, not breastfeeding and mode of delivery. Pharmacological prevention of MTCT is not possible at the present time because both pegylated interferon alfa and ribavirin are contraindicated for use in pregnancy and during the neonatal period. However, this may change with the recent introduction of direct acting antiviral agents. This review summarizes what is currently known about HCV infection during pregnancy and childhood. Particular emphasis is placed on how pregnancy-associated immune modulation may influence the progression of HCV disease and impact MTCT, and on the differential evolution of perinatally acquired HCV infection in children. Taken together, these developments provide insights into the pathogenesis of hepatitis C and may inform strategies to prevent the transmission of HCV from mother to child. Viruses 2012-12-06 4 12 Review 10.3390/v4123531 3531 3550 1999-4915 2012-12-06 doi: 10.3390/v4123531 Armelle Le Campion Ariane Larouche Sébastien Fauteux-Daniel Hugo Soudeyns http://www.mdpi.com/1999-4915/4/12/3511 The filovirus plaque assay serves as the assay of choice to measure infectious virus in a cell culture, blood, or homogenized tissue sample. It has been in use for more than 30 years and is the generally accepted assay used to titrate virus in samples from animals treated with a potential antiviral therapeutic or vaccine. As these animal studies are required for the development of vaccines and therapeutics under the FDA Animal Rule, it is essential to have a standardized assay to compare their efficacies against the various filoviruses. Here, we present an evaluation of the conditions under which the filovirus plaque assay performs best for the Ebola virus Kikwit variant and the Angola variant of Marburg virus. The indicator cell type and source, inoculum volumes, length of incubation and general features of filovirus biology as visualized in the assay are addressed in terms of the impact on the sample viral titer calculations. These optimization studies have resulted in a plaque assay protocol which can be used for preclinical studies, and as a standardized protocol for use across institutions, to aid in data comparison. This protocol will be validated for use in GLP studies supporting advanced development of filovirus therapeutics and vaccines. Viruses 2012-12-06 4 12 Article 10.3390/v4123511 3511 3530 1999-4915 2012-12-06 doi: 10.3390/v4123511 Amy Shurtleff Julia Biggins Ashley Keeney Elizabeth Zumbrun Holly Bloomfield Ana Kuehne Jennifer Audet Kendra Alfson Anthony Griffiths Gene Olinger Sina Bavari http://www.mdpi.com/1999-4915/4/12/3494 Pneumonia Virus of Mice (PVM) is related to the human and bovine respiratory syncytial virus (RSV) pathogens, and has been used to study respiratory virus replication and the ensuing inflammatory response as a component of a natural host—pathogen relationship. As such, PVM infection in mice reproduces many of the clinical and pathologic features of the more severe forms of RSV infection in human infants. Here we review some of the most recent findings on the basic biology of PVM infection and its use as a model of disease, most notably for explorations of virus infection and allergic airways disease, for vaccine evaluation, and for the development of immunomodulatory strategies for acute respiratory virus infection. Viruses 2012-12-05 4 12 Review 10.3390/v4123494 3494 3510 1999-4915 2012-12-05 doi: 10.3390/v4123494 Kimberly Dyer Katia Garcia-Crespo Stephanie Glineur Joseph Domachowske Helene Rosenberg http://www.mdpi.com/1999-4915/4/12/3468 Countering aerosolized filovirus infection is a major priority of biodefense research.  Aerosol models of filovirus infection have been developed in knock-out mice, guinea pigs and non-human primates; however, filovirus infection of immunocompetent mice by the aerosol route has not been reported.  A murine model of aerosolized filovirus infection in mice should be useful for screening vaccine candidates and therapies.  In this study, various strains of wild-type and immunocompromised mice were exposed to aerosolized wild-type (WT) or mouse-adapted (MA) Ebola virus (EBOV).  Upon exposure to aerosolized WT-EBOV, BALB/c, C57BL/6 (B6), and DBA/2 (D2) mice were unaffected, but 100% of severe combined immunodeficiency (SCID) and 90% of signal transducers and activators of transcription (Stat1) knock-out (KO) mice became moribund between 7–9 days post-exposure (dpe).  Exposure to MA-EBOV caused 15% body weight loss in BALB/c, but all mice recovered.  In contrast, 10–30% lethality was observed in B6 and D2 mice exposed to aerosolized MA-EBOV, and 100% of SCID, Stat1 KO, interferon (IFN)-γ KO and Perforin KO mice became moribund between 7–14 dpe. In order to identify wild-type, inbred, mouse strains in which exposure to aerosolized MA-EBOV is uniformly lethal, 60 BXD (C57BL/6 crossed with DBA/2) recombinant inbred (RI) and advanced RI (ARI) mouse strains were exposed to aerosolized MA-EBOV, and monitored for disease severity. A complete spectrum of disease severity was observed. All BXD strains lost weight but many recovered. However, infection was uniformly lethal within 7 to 12 days post-exposure in five BXD strains.  Aerosol exposure of these five BXD strains to 10-fold less MA-EBOV resulted in lethality ranging from 0% in two strains to 90–100% lethality in two strains.  Analysis of post-mortem tissue from BXD strains that became moribund and were euthanized at the lower dose of MA-EBOV, showed liver damage in all mice as well as lung lesions in two of the three strains.  The two BXD strains that exhibited 90–100% mortality, even at a low dose of airborne MA-EBOV will be useful mouse models for testing vaccines and therapies. Additionally, since disease susceptibility is affected by complex genetic traits, a systems genetics approach was used to identify preliminary gene loci modulating disease severity among the panel BXD strains. Preliminary quantitative trait loci (QTLs) were identified that are likely to harbor genes involved in modulating differential susceptibility to Ebola infection. Viruses 2012-12-03 4 12 Article 10.3390/v4123468 3468 3493 1999-4915 2012-12-03 doi: 10.3390/v4123468 Elizabeth Zumbrun Nourtan Abdeltawab Holly Bloomfield Taylor Chance Donald Nichols Paige Harrison Malak Kotb Aysegul Nalca http://www.mdpi.com/1999-4915/4/12/3452 The bovine respiratory syncytial virus (BRSV) is an enveloped, negative sense, single-stranded RNA virus belonging to the pneumovirus genus within the family Paramyxoviridae. BRSV has been recognized as a major cause of respiratory disease in young calves since the early 1970s. The analysis of BRSV infection was originally hampered by its characteristic lability and poor growth in vitro. However, the advent of numerous immunological and molecular methods has facilitated the study of BRSV enormously. The knowledge gained from these studies has also provided the opportunity to develop safe, stable, attenuated virus vaccine candidates. Nonetheless, many aspects of the epidemiology, molecular epidemiology and evolution of the virus are still not fully understood. The natural course of infection is rather complex and further complicates diagnosis, treatment and the implementation of preventive measures aimed to control the disease. Therefore, understanding the mechanisms by which BRSV is able to establish infection is needed to prevent viral and disease spread. This review discusses important information regarding the epidemiology and molecular epidemiology of BRSV worldwide, and it highlights the importance of viral evolution in virus transmission. Viruses 2012-11-30 4 12 Review 10.3390/v4123452 3452 3467 1999-4915 2012-11-30 doi: 10.3390/v4123452 Rosa Sarmiento-Silva Yuko Nakamura-Lopez Gilberto Vaughan http://www.mdpi.com/1999-4915/4/12/3440 Coronaviruses are single stranded, positive sense RNA viruses, which induce the rearrangement of cellular membranes upon infection of a host cell. This provides the virus with a platform for the assembly of viral replication complexes, improving efficiency of RNA synthesis. The membranes observed in coronavirus infected cells include double membrane vesicles. By nature of their double membrane, these vesicles resemble cellular autophagosomes, generated during the cellular autophagy pathway. In addition, coronavirus infection has been demonstrated to induce autophagy. Here we review current knowledge of coronavirus induced membrane rearrangements and the involvement of autophagy or autophagy protein microtubule associated protein 1B light chain 3 (LC3) in coronavirus replication. Viruses 2012-11-30 4 12 Review 10.3390/v4123440 3440 3451 1999-4915 2012-11-30 doi: 10.3390/v4123440 Helena Maier Paul Britton http://www.mdpi.com/1999-4915/4/12/3420 The ubiquitous Epstein-Barr virus (EBV) is associated with several human tumors, which include lymphoid and epithelial malignancies. It is known that EBV persistently infects the memory B cell pool of healthy individuals by activating growth and survival signaling pathways that can contribute to B cell lymphomagenesis.  Although the monoclonal proliferation of EBV-infected cells can be observed in epithelial tumors, such as nasopharyngeal carcinoma and EBV-associated gastric carcinoma, the precise role of EBV in the carcinogenic progress is not fully understood. This review features characteristics and current understanding of EBV-associated gastric carcinoma. EBV-associated gastric carcinoma comprises almost 10% of all gastric carcinoma cases and expresses restricted EBV latent genes (Latency I). Firstly, definition, epidemiology, and clinical features are discussed. Then, the route of infection and carcinogenic role of viral genes are presented.  Of particular interest, the association with frequent genomic CpG methylation and role of miRNA for carcinogenesis are topically discussed. Finally, the possibility of therapies targeting EBV-associated gastric carcinoma is proposed. Viruses 2012-11-29 4 12 Review 10.3390/v4123420 3420 3439 1999-4915 2012-11-29 doi: 10.3390/v4123420 Hisashi Iizasa Asuka Nanbo Jun Nishikawa Masahisa Jinushi Hironori Yoshiyama http://www.mdpi.com/1999-4915/4/12/3389 Prion diseases or transmissible spongiform encephalopathies are a unique category of infectious protein-misfolding neurodegenerative disorders. Hypothesized to be caused by misfolding of the cellular prion protein these disorders possess an infectious quality that thrives in immune-competent hosts. While much has been discovered about the routing and critical components involved in the peripheral pathogenesis of these agents there are still many aspects to be discovered. Research into this area has been extensive as it represents a major target for therapeutic intervention within this group of diseases. The main focus of pathological damage in these diseases occurs within the central nervous system. Cells of the innate immune system have been proven to be critical players in the initial pathogenesis of prion disease, and may have a role in the pathological progression of disease. Understanding how prions interact with the host innate immune system may provide us with natural pathways and mechanisms to combat these diseases prior to their neuroinvasive stage. We present here a review of the current knowledge regarding the role of the innate immune system in prion pathogenesis. Viruses 2012-11-28 4 12 Review 10.3390/v4123389 3389 3419 1999-4915 2012-11-28 doi: 10.3390/v4123389 Barry Bradford Neil Mabbott http://www.mdpi.com/1999-4915/4/12/3363 Avian influenza virus infections in the human population are rare due to their inefficient direct human-to-human transmission. However, when humans are infected, a strong inflammatory response is usually induced, characterized by elevated levels of cytokines and chemokines in serum, believed to be important in the severe pathogenesis that develops in a high proportion of these patients. Extensive research has been performed to understand the molecular viral mechanisms involved in the H5N1 pathogenesis in humans, providing interesting insights about the virus-host interaction and the regulation of the innate immune response by these highly pathogenic viruses. In this review we summarize and discuss the most important findings in this field, focusing mainly on H5N1 virulence factors and their impact on the modulation of the innate immunity in humans. Viruses 2012-11-26 4 12 Review 10.3390/v4123363 3363 3388 1999-4915 2012-11-26 doi: 10.3390/v4123363 Irene Ramos Ana Fernandez-Sesma http://www.mdpi.com/1999-4915/4/12/3336 Filoviruses cause severe hemorrhagic fever in humans with high case-fatality rates. The cellular factors exploited by filoviruses for their spread constitute potential targets for intervention, but are incompletely defined. The viral glycoprotein (GP) mediates filovirus entry into host cells. Recent studies revealed important insights into the host cell molecules engaged by GP for cellular entry. The binding of GP to cellular lectins was found to concentrate virions onto susceptible cells and might contribute to the early and sustained infection of macrophages and dendritic cells, important viral targets. Tyrosine kinase receptors were shown to promote macropinocytic uptake of filoviruses into a subset of susceptible cells without binding to GP, while interactions between GP and human T cell Ig mucin 1 (TIM-1) might contribute to filovirus infection of mucosal epithelial cells. Moreover, GP engagement of the cholesterol transporter Niemann-Pick C1 was demonstrated to be essential for GP-mediated fusion of the viral envelope with a host cell membrane. Finally, mutagenic and structural analyses defined GP domains which interact with these host cell factors. Here, we will review the recent progress in elucidating the molecular interactions underlying filovirus entry and discuss their implications for our understanding of the viral cell tropism. Viruses 2012-11-26 4 12 Review 10.3390/v4123336 3336 3362 1999-4915 2012-11-26 doi: 10.3390/v4123336 Heike Hofmann-Winkler Franziska Kaup Stefan Pöhlmann