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  • 刊频: Continuously updated 2017-
  • NLM标题: J Gen Virol
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  • 机译 关于将人类鼻病毒AB和C分为基因型指定类型的提案
    摘要:Human rhinoviruses (HRVs) frequently cause mild upper respiratory tract infections and more severe disease manifestations such as bronchiolitis and asthma exacerbations. HRV is classified into three species within the genus Enterovirus of the family Picornaviridae. HRV species A and B contain 75 and 25 serotypes identified by cross-neutralization assays, although the use of such assays for routine HRV typing is hampered by the large number of serotypes, replacement of virus isolation by molecular methods in HRV diagnosis and the poor or absent replication of HRV species C in cell culture. To address these problems, we propose an alternative, genotypic classification of HRV-based genetic relatedness analogous to that used for enteroviruses. Nucleotide distances between 384 complete VP1 sequences of currently assigned HRV (sero)types identified divergence thresholds of 13, 12 and 13 % for species A, B and C, respectively, that divided inter- and intra-type comparisons. These were paralleled by 10, 9.5 and 10 % thresholds in the larger dataset of >3800 VP4 region sequences. Assignments based on VP1 sequences led to minor revisions of existing type designations (such as the reclassification of serotype pairs, e.g. A8/A95 and A29/A44, as single serotypes) and the designation of new HRV types A101–106, B101–103 and C34–C51. A protocol for assignment and numbering of new HRV types using VP1 sequences and the restriction of VP4 sequence comparisons to type identification and provisional type assignments is proposed. Genotypic assignment and identification of HRV types will be of considerable value in the future investigation of type-associated differences in disease outcomes, transmission and epidemiology.
  • 机译 MERS冠状病毒复制诱导严重的体外细胞病理学并被环孢菌素A或干扰素-α治疗强烈抑制
    摘要:Coronavirus (CoV) infections are commonly associated with respiratory and enteric disease in humans and animals. The 2003 outbreak of severe acute respiratory syndrome (SARS) highlighted the potentially lethal consequences of CoV-induced disease in humans. In 2012, a novel CoV (Middle East Respiratory Syndrome coronavirus; MERS-CoV) emerged, causing 49 human cases thus far, of which 23 had a fatal outcome. In this study, we characterized MERS-CoV replication and cytotoxicity in human and monkey cell lines. Electron microscopy of infected Vero cells revealed extensive membrane rearrangements, including the formation of double-membrane vesicles and convoluted membranes, which have been implicated previously in the RNA synthesis of SARS-CoV and other CoVs. Following infection, we observed rapidly increasing viral RNA synthesis and release of high titres of infectious progeny, followed by a pronounced cytopathology. These characteristics were used to develop an assay for antiviral compound screening in 96-well format, which was used to identify cyclosporin A as an inhibitor of MERS-CoV replication in cell culture. Furthermore, MERS-CoV was found to be 50–100 times more sensitive to alpha interferon (IFN-α) treatment than SARS-CoV, an observation that may have important implications for the treatment of MERS-CoV-infected patients. MERS-CoV infection did not prevent the IFN-induced nuclear translocation of phosphorylated STAT1, in contrast to infection with SARS-CoV where this block inhibits the expression of antiviral genes. These findings highlight relevant differences between these distantly related zoonotic CoVs in terms of their interaction with and evasion of the cellular innate immune response.
  • 机译 Toll样受体9基因启动子区多态性与宫颈人乳头瘤病毒感染
    摘要:Polymorphism in the Toll-like receptor (TLR) 9 gene has been shown to have a significant role in some diseases; however, little is known about its possible role in the natural history of human papillomavirus (HPV) infections. We investigated the association between a single-nucleotide polymorphism (SNP) (rs5743836) in the promoter region of TLR9 (T1237C) and type-specific HPV infections. Specimens were derived from a cohort of 2462 women enrolled in the Ludwig–McGill Cohort Study. We randomly selected 500 women who had a cervical HPV infection detected at least once during the study as cases. We defined two control groups: (i) a random sample of 300 women who always tested HPV negative, and (ii) a sample of 234 women who were always HPV negative but had a minimum of ten visits during the study. TLR9 genotyping was performed using bidirectional PCR amplification of specific alleles. Irrespective of group, the WT homozygous TLR9 genotype (TT) was the most common form, followed by the heterozygous (TC) and the mutant homozygous (CC) forms. There were no consistent associations between polymorphism and infection risk, either overall or by type or species. Likewise, there were no consistently significant associations between polymorphism and HPV clearance or persistence. We concluded that this polymorphism in the promoter region of TLR9 gene does not seem to have a mediating role in the natural history of the HPV infection.
  • 机译 火鸡中新的小核糖核酸病毒(Meleagris gallopavo)的遗传特征不同于火鸡的鸡微小病毒和Megriviruses并且与禽肝炎病毒属成员密切相关
    摘要:This study reports the metagenomic detection and complete genome characterization of a novel turkey picornavirus from faecal samples of healthy (1/3) and affected (6/8) commercial turkeys with enteric and/or stunting syndrome in Hungary. The virus was detected at seven of the eight farms examined. The turkey/M176-TuASV/2011/HUN genome () was genetically different from the currently known picornaviruses of turkey origin (megriviruses and galliviruses), and showed distant phylogenetic relationship and common genomic features (e.g. uncleaved VP0 and three predicted and unrelated 2A polypeptides) to duck hepatitis A virus (DHAV) of the genus Avihepatovirus. The complete genome analysis revealed multiple distinct genome features like the presence of two in-tandem aphthovirus 2A-like sequence repeats with DxExNPG/P ‘ribosome-skipping’ sites (76 %, 23/30 amino acids identical), with the first aphthovirus 2A-like sequence being located at the end of the VP1 capsid protein (VP1/2A1 ‘ribosome-skipping’ site). The phylogenetic analyses, low sequence identity (33, 32 and 36 % amino acid identity in P1, P2 and P3 regions) to DHAV, and the type II-like internal ribosome entry site suggests that this turkey picornavirus is related to, but distinct from the genus Avihepatovirus and it could be the founding member of a novel Avihepatovirus sister-clade genus. This is the third, taxonomically highly distinct picornavirus clade identified from turkeys exhibiting varied symptoms.
  • 机译 必需的RNA元件位于戊型肝炎病毒ORF2的中央区域
    摘要:Hepatitis E virus (genus Hepevirus, family Hepeviridae) is one of the most important causes of acute hepatitis in adults, particularly among pregnant women, throughout Asia and Africa where mortality rates can be 20–30 %. Hepatitis E virus has a single-stranded positive-sense RNA genome that contains three translated ORFs. The two 3′ ORFs are translated from a subgenomic RNA. Functional RNA elements have been identified in and adjacent to the genomic 5′ and 3′ UTRs and in and around the intergenic region. Here we describe an additional RNA element that is located in a central region of ORF2. The RNA element is predicted to fold into two highly conserved stem–loop structures, ISL1 and ISL2. Mutations that disrupt the predicted structures, without altering the encoded amino acid sequence, result in a drastic reduction in capsid protein synthesis. This indicates that the RNA element plays an important role in one of the early steps of virus replication. The structures were further investigated using a replicon that expresses Gaussia luciferase in place of the capsid protein. Single mutations in ISL2 severely reduced luciferase expression, but a pair of compensatory mutations that were predicted to restore the ISL2 structure, restored luciferase expression to near-WT levels, thus lending experimental support to the predicted structure. Nonetheless the precise role of the ISL1+ISL2 element remains unknown.
  • 机译 轮状病毒感染细胞的脂质组分析证实脂质滴与病毒质的紧密相互作用
    摘要:Rotaviruses (RVs) cause acute gastroenteritis in infants and young children, and are globally distributed. Within the infected host cell, RVs establish replication complexes in viroplasms (‘viral factories’) to which lipid droplet organelles are recruited. To further understand this recently discovered phenomenon, the lipidomes of RV-infected and uninfected MA104 cells were investigated. Cell lysates were subjected to equilibrium ultracentrifugation through iodixanol gradients. Fourteen different classes of lipids were differentiated by mass spectrometry. The concentrations of virtually all lipids were elevated in RV-infected cells. Fractions of low density (1.11–1.15 g ml−1), in which peaks of the RV dsRNA genome and lipid droplet- and viroplasm-associated proteins were observed, contained increased amounts of lipids typically found concentrated in the cellular organelle lipid droplets, confirming the close interaction of lipid droplets with viroplasms. A decrease in the ratio of the amounts of surface to internal components of lipid droplets upon RV infection suggested that the lipid droplet–viroplasm complexes became enlarged.
  • 机译 牛痘病毒蛋白K7是一种毒力因子可改变对感染的急性免疫反应
    摘要:Vaccinia virus (VACV) encodes many proteins that antagonize the innate immune system including a family of intracellular proteins with a B-cell lymphoma (Bcl)-2-like structure. One of these Bcl-2 proteins called K7 binds Toll-like receptor-adaptor proteins and the DEAD-box RNA helicase DDX3 and thereby inhibits the activation of NF-κB and interferon regulatory factor 3. However, the contribution of K7 to virus virulence is not known. Here a VACV lacking the K7R gene (vΔK7) was constructed and compared with control viruses that included a plaque purified wt (vK7), a revertant with the K7R gene reinserted (vK7-rev) and a frame-shifted virus in which the translational initiation codon was mutated to prevent K7 protein expression (vK7-fs). Data presented show that loss of K7 does not affect virus replication in cell culture or in vivo; however, viruses lacking the K7 protein were less virulent than controls in murine intradermal (i.d.) and intranasal (i.n.) infection models and there was an altered acute immune response to infection. In the i.d. model, vΔK7 induced smaller lesions than controls, and after i.n. infection vΔK7 induced a reduced weight loss and signs of illness, and more rapid clearance of virus from infected tissue. Concomitantly, the intrapulmonary innate immune response to infection with vΔK7 showed increased infiltration of NK cells and CD8+ T-cells, enhanced MHC class II expression by macrophages, and enhanced cytolysis of target cells by NK cells and VACV-specific CD8+ T-cells. Thus protein K7 is a virulence factor that affects the acute immune response to infection.
  • 机译 表达荧光标记蛋白的传染性重组口蹄疫病毒
    摘要:Foot-and-mouth disease virus (FMDV) is one of the most extensively studied animal pathogens because it remains a major threat to livestock economies worldwide. However, the dynamics of FMDV infection are still poorly understood. The application of reverse genetics provides the opportunity to generate molecular tools to further dissect the FMDV life cycle. Here, we have used reverse genetics to determine the capsid packaging limitations for a selected insertion site in the FMDV genome. We show that exogenous RNA up to a defined length can be stably introduced into the FMDV genome, whereas larger insertions are excised by recombination events. This led us to construct a recombinant FMDV expressing the fluorescent marker protein, termed iLOV. Characterization of infectious iLOV-FMDV showed the virus has a plaque morphology and rate of growth similar to the parental virus. In addition, we show that cells infected with iLOV-FMDV are easily differentiated by flow cytometry using the inherent fluorescence of iLOV and that cells infected with iLOV-FMDV can be monitored in real-time with fluorescence microscopy. iLOV-FMDV therefore offers a unique tool to characterize FMDV infection in vitro, and its applications for in vivo studies are discussed.
  • 机译 在马拉维超过十年(1997-2007年)检测到的具有保守基因型星座的G8轮状病毒显示出人类共同传播的菌株之间频繁的基因重排
    摘要:Rotavirus A, the most common cause of severe diarrhoea in children worldwide, occurs in five major VP7 (G) and VP4 (P) genotype combinations, comprising G1P[8], G2P[4], G3P[8], G4P[8] and G9P[8]. However, G8, a common bovine rotavirus genotype, has been reported frequently among children in African countries. Surveillance of rotavirus gastroenteritis conducted in a sentinel hospital in Blantyre, Malawi between 1997 and 2007 provided a rare opportunity to examine the whole genotype constellation of G8 strains and their evolution over time. A sample of 27 (9.0 %) of 299 G8 strains was selected to represent each surveillance year and a range of P genotypes, which shifted in predominance from P[6] to P[4] and P[8] during the study period. Following cell culture adaptation, whole genome sequencing demonstrated that the genetic background of 26 strains possessed the DS-1 genotype constellation. A single G8P[6] strain was a reassortant in which both NSP2 and NSP5 genes from strains with the Wa genotype constellation had been inserted into a strain with the DS-1 genotype background. Phylogenetic analysis suggested frequent reassortment among co-circulating strains with the DS-1 genotype constellation. Little evidence was identified to suggest the introduction of contemporary bovine rotavirus genes into any of the 27 G8 strains examined. In conclusion, Malawian G8 strains are closely related to other human strains with the DS-1 genotype constellation. They have evolved over the last decade through genetic reassortment with other human rotaviruses, changing their VP4 genotypes while maintaining a conserved genotype constellation for the remaining structural and non-structural proteins.
  • 机译 新城疫病毒融合蛋白和血凝素神经氨酸酶蛋白有助于其巨噬细胞宿主范围
    摘要:The fusion (F) and haemagglutinin-neuraminidase (HN) proteins of Newcastle disease virus (NDV) are multifunctional proteins that play critical roles during infection. Here, we assessed the ability of NDV to replicate in macrophages and investigated the contribution of the F and HN proteins to NDV infection/replication in these cells. Results of our study revealed that, while presenting similar replication kinetics in a fibroblast cell line (DF1) or in primary non-adherent splenocytes, the NDV strain CA02 replicates better in macrophages (HD11 and primary adherent splenocytes) than the NDV strain Anhinga/93. Notably, exchange of the HN or both F and HN genes of NDV Anhinga/93 by the corresponding genes from NDV CA02 markedly improved the ability of the chimeric viruses to replicate in macrophages. These results indicate that the F and HN proteins are determinants of NDV macrophage host range. This represents the first description of productive NDV infection in macrophages.
  • 机译 沙眼病毒反向遗传学用于疫苗开发
    摘要:Arenaviruses are important human pathogens with no Food and Drug Administration (FDA)-licensed vaccines available and current antiviral therapy being limited to an off-label use of the nucleoside analogue ribavirin of limited prophylactic efficacy. The development of reverse genetics systems represented a major breakthrough in arenavirus research. However, rescue of recombinant arenaviruses using current reverse genetics systems has been restricted to rodent cells. In this study, we describe the rescue of recombinant arenaviruses from human 293T cells and Vero cells, an FDA-approved line for vaccine development. We also describe the generation of novel vectors that mediate synthesis of both negative-sense genome RNA and positive-sense mRNA species of lymphocytic choriomeningitis virus (LCMV) directed by the human RNA polymerases I and II, respectively, within the same plasmid. This approach reduces by half the number of vectors required for arenavirus rescue, which could facilitate virus rescue in cell lines approved for human vaccine production but that cannot be transfected at high efficiencies. We have shown the feasibility of this approach by rescuing both the Old World prototypic arenavirus LCMV and the live-attenuated vaccine Candid#1 strain of the New World arenavirus Junín. Moreover, we show the feasibility of using these novel strategies for efficient rescue of recombinant tri-segmented both LCMV and Candid#1.
  • 机译 轮状病毒峰值蛋白VP4中的突变降低了胰蛋白酶的敏感性但没有病毒传播
    摘要:Infectious entry of the nonenveloped rotavirus virion requires proteolysis of the spike protein VP4 to mediate conformational changes associated with membrane penetration. We sequenced and characterized an isolate that was cultured in the absence of trypsin and found that it is more resistant to proteolysis than WT virus. A substitution mutation abrogates one of the defined trypsin-cleavage sites, suggesting that blocking proteolysis at this site reduces the overall kinetics of proteolysis. Kinetic analysis of the membrane penetration-associated conformational change indicated that the ‘fold-back’ of the mutant spike protein is slower than that of WT. Despite these apparent biochemical defects, the mutant virus replicates in an identical manner to the WT virus. These findings enhance an understanding of VP4 functions and establish new strategies to interrogate rotavirus cell entry.
  • 机译 脊椎动物减毒的西尼罗河病毒突变体对库塔尔蚊的载体能力具有不同影响
    摘要:Previous mutational analyses of naturally occurring West Nile virus (WNV) strains and engineered mutant WNV strains have identified locations in the viral genome that can have profound phenotypic effect on viral infectivity, temperature sensitivity and neuroinvasiveness. We chose six mutant WNV strains to evaluate for vector competence in the natural WNV vector Culex tarsalis, two of which contain multiple ablations of glycosylation sites in the envelope and NS1 proteins; three of which contain mutations in the NS4B protein and an attenuated natural bird isolate (Bird 1153) harbouring an NS4B mutation. Despite vertebrate attenuation, all NS4B mutant viruses displayed enhanced vector competence by Cx. tarsalis. Non-glycosylated mutant viruses displayed decreased vector competence in Cx. tarsalis mosquitoes, particularly when all three NS1 glycosylation sites were abolished. These results indicate the importance of both the NS4B protein and NS1 glycosylation in the transmission of WNV by a significant mosquito vector.
  • 机译 来自墨西哥的蝙蝠中的冠状病毒
    摘要:Bats are reservoirs for a wide range of human pathogens including Nipah, Hendra, rabies, Ebola, Marburg and severe acute respiratory syndrome coronavirus (CoV). The recent implication of a novel beta (β)-CoV as the cause of fatal respiratory disease in the Middle East emphasizes the importance of surveillance for CoVs that have potential to move from bats into the human population. In a screen of 606 bats from 42 different species in Campeche, Chiapas and Mexico City we identified 13 distinct CoVs. Nine were alpha (α)-CoVs; four were β-CoVs. Twelve were novel. Analyses of these viruses in the context of their hosts and ecological habitat indicated that host species is a strong selective driver in CoV evolution, even in allopatric populations separated by significant geographical distance; and that a single species/genus of bat can contain multiple CoVs. A β-CoV with 96.5 % amino acid identity to the β-CoV associated with human disease in the Middle East was found in a Nyctinomops laticaudatus bat, suggesting that efforts to identify the viral reservoir should include surveillance of the bat families Molossidae/Vespertilionidae, or the closely related Nycteridae/Emballonuridae. While it is important to investigate unknown viral diversity in bats, it is also important to remember that the majority of viruses they carry will not pose any clinical risk, and bats should not be stigmatized ubiquitously as significant threats to public health.
  • 机译 NS5A域II的C端是丙型肝炎病毒基因组复制的关键决定因素但病毒体组装和释放并不需要
    摘要:The NS5A protein of hepatitis C virus (HCV) plays roles in both virus genome replication and the assembly of infectious virus particles. NS5A comprises three domains, separated by low-complexity sequences. Whilst the function of domain I appears to be predominantly involved with genome replication, the roles of domains II and III are less well defined. It has been reported previously that a deletion spanning the majority of domain II but retaining the C-terminal 35 residues had no effect on virus production; however, deletion of the entire domain II eliminated genome replication, pointing to a key role for the C terminus of this domain. Recent work has also highlighted this region as the potential binding site of the host factor cyclophilin A (CypA). To define this requirement for replication in more detail, and to investigate the involvement of CypA, we conducted a mutagenic study of the C-terminal 30 residues of domain II within the context of both the infectious JFH-1 virus and a JFH-1-derived subgenomic replicon. We showed that 12 of these residues were absolutely required for virus genome replication, whilst mutations of the remainder either had no phenotype or exhibited a partial reduction in genome replication. There was an absolute correlation between the datasets for virus and subgenomic replicon, indicating that this region is involved solely in the process of genome replication. Comparison of our data with a previously published analysis of the same region in genotype 1b revealed some important differences between the two genotypes of HCV.
  • 机译 人类鼻病毒2Apro的特异性由四个切割位点残基的组合空间特性决定
    摘要:
  • 机译 具有不同M1蛋白的甲型H1N1 WSN33流感病毒的体内外复制
    摘要:The M1 protein is a major structural protein that has multiple functions in various steps within the life cycle of the influenza A virus (IAV). However, little is currently known about the role of M1 in IAV replication in vivo and the associated pathogenesis. In this study, six isogenic H1N1 WSN33 viruses, constructed to express unique M1 proteins derived from various strains, subtypes or WSN33 itself, were tested to determine in vitro and in vivo functional exchangeability of M1 proteins in the replication and pathogenesis of the WSN33 virus. Despite five chimeric M1 viruses replicating to levels similar to those of the parental WSN33 virus in cell cultures, all M1 chimeras exhibited improved replication and enhanced virulence in mice when compared with the WSN33 virus. Interestingly, M1 proteins derived from swine viruses caused more severe clinical diseases than those from human or quail. These data indicate that the M1 protein is an important determinant of viral replication and pathogenic properties in mice, although the functions of M1 observed in vivo are not adequately reflected in simple infections of cultured cells. Chimeric M1 viruses that are variable in their clinical manifestations described here will aid future understanding of the role of M1 in IAV pathogenesis.
  • 机译 猪瘟病毒(Bunyaviridae)的Salehabad病毒物种复合体的表征
    摘要:Genomic and antigenic characterization of the Salehabad virus, a species of the genus Phlebovirus, and four other unclassified phleboviruses (Arbia, Adria, Arumowot and Odrenisrou) demonstrate a serological and genetic relation to one another and are distinct from the eight other recognized species within the genus Phlebovirus. We propose to incorporate these four unclassified viruses as part of the Salehabad species complex within the genus. The known geographical distribution for the members of this species group includes southern Europe, Central Asia and Africa.
  • 机译 牛痘病毒A34蛋白的运输和稳定性受A33蛋白影响
    摘要:Vaccinia virus (VACV) has two infectious forms called intracellular mature virus and extracellular enveloped virus (EEV). Two of the seven viral proteins in the EEV outer envelope, A33 and A34, are type II membrane glycoproteins that each interact with another EEV protein called B5; however, evidence for direct A33–A34 interaction is lacking. The localization and stability of A34 is affected by B5 and here data are presented showing that A34 is also affected by A33. In the absence of A33, just as without B5, the level, localization and glycosylation profile of A34 was altered. However, the glycosylation profile of A34 without A33 is different to that observed in the absence of B5, and A34 accumulates in the Golgi apparatus rather than in the endoplasmic reticulum. Thus, A34 requires more than one other EEV protein for its processing and cellular transport.
  • 机译 黑猩猩GB病毒C和GB病毒A E2包膜糖蛋白含有抑制人类CD4 + T细胞中人类免疫缺陷病毒1型复制的肽基序
    摘要:GB virus type C (GBV-C) is a lymphotropic virus that can cause persistent infection in humans. GBV-C is not associated with any disease, but is associated with reduced mortality in human immunodeficiency virus type 1 (HIV-1)-infected individuals. Related viruses have been isolated from chimpanzees (GBV-Ccpz) and from New World primates (GB virus type A, GBV-A). These viruses are also capable of establishing persistent infection. We determined the nucleotide sequence encoding the envelope glycoprotein (E2) of two GBV-Ccpz isolates obtained from the sera of captive chimpanzees. The deduced GBV-Ccpz E2 protein differed from human GBV-C by 31 % at the amino acid level. Similar to human GBV-C E2, expression of GBV-Ccpz E2 in a tet-off human CD4+ Jurkat T-cell line significantly inhibited the replication of diverse HIV-1 isolates. This anti-HIV-replication effect of GBV-Ccpz E2 protein was reversed by maintaining cells in doxycycline to reduce E2 expression. Previously, we found a 17 aa region within human GBV-C E2 that was sufficient to inhibit HIV-1. Although GBV-Ccpz E2 differed by 3 aa differences in this region, the chimpanzee GBV-C 17mer E2 peptide inhibited HIV-1 replication. Similarly, the GBV-A peptide that aligns with this GBV-C E2 region inhibited HIV-1 replication despite sharing only 5 aa with the human GBV-C E2 sequence. Thus, despite amino acid differences, the peptide region on both the GBV-Ccpz and the GBV-A E2 protein inhibit HIV-1 replication similar to human GBV-C. Consequently, GBV-Ccpz or GBV-A infection of non-human primates may provide an animal model to study GB virus–HIV interactions.

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