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1.  Identification and characterization of two novel human papillomaviruses (HPVs) by overlapping PCR: HPV102 and HPV106 
The Journal of general virology  2007;88(0 11):2952-2955.
Complete genomes of HPV102 (8078 bp) and HPV106 (8035 bp) were PCR amplified and cloned from cervicovaginal cells of a 49-year-old Hispanic female with reactive changes on her Pap test and a 42-year-old Hispanic female with a Pap test diagnosis of atypical squamous cells of unknown significance (ASCUS), respectively. The nucleotide sequence similarity of the complete L1 open reading frame (ORF) determined that HPV102 and HPV106 are most closely related to HPV83 (84.1 % identity) and HPV90 (83.5 % identity), respectively, placing them in the genital HPV groups, papillomaviruses species α3 and α15. HPV102 and HPV106 contain five early genes (E6, E7, E1, E2, and E4) and two late genes (L2 and L1), and both lack an E5 ORF. On the basis of phylogenetic analyses and available clinical information, these two novel HPV types expand the heterogeneity of HPVs detected in the lower genital tract.
doi:10.1099/vir.0.83178-0
PMCID: PMC3963832  PMID: 17947516
2.  The human immunodeficiency virus type 1 Vpr protein upregulates PVR via activation of the ATR-mediated DNA damage response pathway 
The Journal of general virology  2013;94(0 12):2664-2669.
Viral infection may induce the cell-surface expression of PVR (CD155) that, upon recognition by its cognate activating DNAM-1 receptor present on cytotoxic lymphocytes, may promote antiviral immune responses. Here we show that expression of the human immunodeficiency virus type 1 (HIV-1) Vpr protein in Jurkat T cells increases cell-surface and total PVR levels. Analysis of mutated Vpr variants indicated that Vpr uses the same protein surfaces, and hence probably the same mechanisms, to upregulate PVR and arrest the cell cycle in the G2 phase. Moreover, we found that PVR upregulation by Vpr relied on the ability of the protein to activate the ATR kinase that triggers the DNA damage response pathway and G2 arrest. Finally, we showed that Vpr contributes to PVR up-modulation in HIV-infected CD4+ T lymphocytes and inhibits the PVR downregulating activity of the viral Nef protein.
doi:10.1099/vir.0.055541-0
PMCID: PMC3955189  PMID: 24045107 CAMSID: cams4193
3.  Generation and genetic stability of tick-borne encephalitis virus mutants dependent on processing by the foot-and-mouth disease virus 3C protease 
The Journal of general virology  2011;93(0 3):504-515.
Mature protein C of tick-borne encephalitis virus (TBEV) is cleaved from the polyprotein precursor by the viral NS2B/3 protease (NS2B/3pro). We showed previously that replacement of the NS2B/3pro cleavage site at the C terminus of protein C by the foot-and-mouth disease virus (FMDV) 2A StopGo sequence leads to the production of infectious virions. Here, we show that infectious virions can also be produced from a TBEV mutant bearing an inactivated 2A sequence through the expression of the FMDV 3C protease (3Cpro) either in cis or in trans (from a TBEV replicon). Cleavage at the C terminus of protein C depended on the catalytic activity of 3Cpro as well as on the presence of an optimized 3Cpro cleavage site. Passage of the TBEV mutants bearing a 3Cpro cleavage site either in the absence of 3Cpro or in the presence of a catalytically inactive 3Cpro led to the appearance of revertants in which protein C cleavage by NS2B/3pro had been regained. In three different revertants, a cleavage site for NS2B/3pro, namely RR*C, was now present, leading to an elongated protein C. Furthermore, two revertants acquired additional mutations in the C terminus of protein C, eliminating two basic residues. Although these latter mutants showed wild-type levels of early RNA synthesis, their foci were smaller and an accumulation of protein C in the cytoplasm was observed. These findings suggest a role of the positive charge of the C terminus of protein C for budding of the nucleocapsid and further support the notion that TBEV protein C is a multifunctional protein.
doi:10.1099/vir.0.038398-0
PMCID: PMC3918513  PMID: 22131310
4.  Serological relationship between cutaneous human papillomavirus types 5, 8 and 92 
The Journal of general virology  2009;90(0 1):136-143.
Evidence of a possible association of cutaneous human papillomavirus (HPV) types, especially members of the genus Betapapillomavirus, and the development of non-melanoma skin cancer (NMSC) is accumulating. Vaccination with virus-like particles (VLPs) consisting of self-assembled L1, the major capsid protein, has been introduced to control anogenital HPV infection. This study examined the serological relationship between betapapillomavirus (β-PV) types 5 and 8 and the new type HPV-92, which has recently been isolated from a basal cell carcinoma containing a high number of viral genomes. Following expression by recombinant baculoviruses, the L1 protein of HPV-92 self-assembled into VLPs that elicited high-titre antibodies after immunization, similar to VLPs from HPV-5 and -8. Haemagglutination inhibition (HAI) assays were used as a surrogate method for the detection of virion-neutralizing antibodies, which correlates with protection from infection. Antisera raised against HPV-5 and -8 VLPs displayed HAI activity not only against the homologous type, but also against heterologous HPV types 5, 8 and 92, whereas HAI activity of antisera against HPV-92 VLP was restricted to the homologous type. The results of neutralization assays using HPV-5 pseudovirions were consistent with those from HAI assays. Cross-neutralizing immune responses by VLP vaccination against heterologous HPV types may provide broader protection against the multiplicity of HPV types detected in NMSC. If a close link to HPV infection can be conclusively established, these results may provide a basis for further evaluation of VLPs of β-PVs as candidates for a prophylactic skin-type HPV vaccine, aimed at reducing the incidence of NMSC.
doi:10.1099/vir.0.006189-0
PMCID: PMC3795330  PMID: 19088282
5.  Chimeric papillomavirus-like particles expressing a foreign epitope on capsid surface loops 
The Journal of general virology  2001;82(0 11):2799-2804.
Neutralization capsid epitopes are important determinants for antibody-mediated immune protection against papillomavirus (PV) infection and induced disease. Chimeric L1 major capsid proteins of the human PV type 16 (HPV-16) and the bovine PV type 1 (BPV-1) with a foreign peptide incorporated into several capsid surface loops self-assembled into pentamers or virus-like particles (VLP). Binding patterns of neutralizing monoclonal antibodies (MAb) and immunization of mice confirmed (i) that regions around aa 282–286 and 351–355 contribute to neutralization epitopes and identified the latter region as an immunodominant site and (ii) that placing a foreign peptide in the context of an assembled structure markedly enhanced its immunogenicity. Pentamers disassembled from wild-type HPV-16 and BPV-1 VLPs displayed some of the neutralization epitopes that were detected on fully assembled VLPs, but were deficient for binding a subset of neutralizing MAb that inhibit cell attachment.
PMCID: PMC3795388  PMID: 11602792
6.  Mutagenesis of the murine cytomegalovirus M56 terminase gene 
The Journal of general virology  2008;89(0 11):2864-2868.
SUMMARY
The M56 gene encodes M56, one of three proteins comprising the murine cytomegalovirus (MCMV) terminase, an enzyme that packages DNA into capsids and cleaves the DNA into progeny genomes. Deletion of M56 from a bacterial artificial chromosome (BAC) clone of the MCMV genome was lethal, as the mutant BAC failed to reconstitute infectious virus. Reintroduction of M56 at an ectopic locus complemented the deletion, allowing reconstitution of a virus that replicated with wild type efficiency. Reintroduction of M56 sequences encoding an N-terminal epitope fusion was lethal, as was a mutation targeting a region in M56 implicated as an ATPase active site. In contrast, a frame shift mutation in M56a, an open reading frame that overlaps M56, had no effect on viral replication. We conclude that the protein product of M56a is dispensable while M56 residues comprising the proposed ATPase active site are critical for terminase function and viral replication.
doi:10.1099/vir.0.2008/003137-0
PMCID: PMC3771075  PMID: 18931084
7.  Measurement of membrane fusion activity from viral membrane fusion proteins based on a fusion-dependent promoter induction system in insect cells 
The Journal of general virology  2001;82(0 10):2519-2529.
Summary
A number of viral membrane fusion proteins can be expressed alone on the surface of host cells, then triggered to induce cell-to-cell fusion or syncytium formation. Although rapid and easily observed, syncytium formation is not easily quantified and differences in fusion activity are not easily distinguished or measured. To address this problem, we developed a rapid and quantitative cell-to-cell fusion system that is useful for comparative analysis and may be suitable for high throughput screening. In this system, expression of a reporter protein, the enhanced green fluorescent protein (EGFP), is dependent on cell-to-cell fusion. Spodoptera frugiperda (Sf9) insect cells expressing a chimeric Lac Repressor-IE1 protein were fused to Sf9 cells containing an EGFP reporter construct under the control of a responsive lac operator containing promoter. Membrane fusion efficiency was measured from the resulting EGFP fluorescence activity. Sf9 cells expressing the Orgyia pseudotsugata Multicapsid Nucleopolyhedrovirus (OpMNPV) GP64 envelope fusion protein were used as a model to test this fusion assay. Subtle changes in fusion activities of GP64 proteins containing single amino acid substitutions in a putative membrane fusion domain were distinguished, and decreases in EGFP fluorescence corresponded to decreases in the hydrophobicity in the small putative membrane fusion domain.
PMCID: PMC3754798  PMID: 11562545
8.  [No title available] 
PMCID: PMC3646623  PMID: 18272766
9.  Development of a reverse genetics system for murine norovirus 3; long-term persistence occurs in the caecum and colon 
The Journal of general virology  2012;93(Pt 7):1432-1441.
SUMMARY
Human noroviruses (HuNoV) are a major cause of viral gastroenteritis worldwide yet due to the inability to propagate HuNoV in cell culture, murine norovirus (MNV) is typically used as a surrogate to study norovirus biology. MNV-3 represents an attractive strain to study norovirus infections in vivo since it establishes persistence in wild type mice, yet causes symptoms resembling gastroenteritis in immune compromised STAT1-/- mice. The lack of reverse genetics approaches to recover genetically defined MNV-3 has limited further studies on the identification of viral sequences that contribute to persistence. Here we report the establishment of a combined DNA based reverse genetics and mouse model system to study persistent MNV-3 infections in wild type (C57BL/6) mice. Viral RNA and infectious virus were detected in the faeces for at least 56 days after inoculation. Strikingly, the highest concentrations of viral RNA during persistence were detected in the caecum and colon, suggesting that viral persistence is maintained in these tissues. Possible adaptive changes arising during persistence in vivo appeared to accumulate in the minor capsid protein (VP2) and the viral polymerase (NS7), in contrast with adaptive mutations selected during cell culture passages in RAW264.7 cells that appear in the major capsid protein (VP1) and non-structural protein NS4. This system provides an attractive model that can be readily used to identify viral sequences that contribute to persistence in an immunocompetent host and to more acute infection in an immunocompromised host, providing new insights into the biology of norovirus infections.
doi:10.1099/vir.0.042176-0
PMCID: PMC3541523  PMID: 22495235
10.  Punique virus, a novel phlebovirus, related to sandfly fever Naples virus, isolated from sandflies collected in Tunisia 
The Journal of general virology  2010;91(Pt 5):1275-1283.
Sandflies are widely distributed around the Mediterranean Basin. Therefore, human populations in this area are potentially exposed to sandfly-transmitted diseases, including those caused by phleboviruses. Whilst there are substantial data in countries located in the northern part of the Mediterranean basin, few data are available for North Africa. In this study, a total of 1489 sandflies were collected in 2008 in Tunisia from two sites, bioclimatically distinct, located 235 km apart, and identified morphologically. Sandfly species comprised Phlebotomus perniciosus (52.2 %), Phlebotomus longicuspis (30.1 %), Phlebotomus papatasi (12 .0%), Phlebotomus perfiliewi (4.6 %), Phlebotomus langeroni (0.4 %) and Sergentomyia minuta (0.5 %). PCR screening, using generic primers for the genus Phlebovirus, resulted in the detection of ten positive pools. Sequence analysis revealed that two pools contained viral RNA corresponding to a novel virus closely related to sandfly fever Naples virus. Virus isolation in Vero cells was achieved from one pool. Genetic and phylogenetic characterization based on sequences in the three genomic segments showed that it was a novel virus distinct from other recognized members of the species. This novel virus was provisionally named Punique virus. Viral sequences in the polymerase gene corresponding to another phlebovirus closely related to but distinct from sandfly fever Sicilian virus were obtained from the eight remaining positive pools.
doi:10.1099/vir.0.019240-0
PMCID: PMC3496376  PMID: 20089800
11.  Development of a reverse-genetics system for murine norovirus 3: long-term persistence occurs in the caecum and colon 
The Journal of General Virology  2012;93(Pt 7):1432-1441.
Human noroviruses (HuNoV) are a major cause of viral gastroenteritis worldwide, yet, due to the inability to propagate HuNoV in cell culture, murine norovirus (MNV) is typically used as a surrogate to study norovirus biology. MNV-3 represents an attractive strain to study norovirus infections in vivo because it establishes persistence in wild-type mice, yet causes symptoms resembling gastroenteritis in immune-compromised STAT1−/− mice. The lack of reverse-genetics approaches to recover genetically defined MNV-3 has limited further studies on the identification of viral sequences that contribute to persistence. Here we report the establishment of a combined DNA-based reverse-genetics and mouse-model system to study persistent MNV-3 infections in wild-type (C57BL/6) mice. Viral RNA and infectious virus were detected in faeces for at least 56 days after inoculation. Strikingly, the highest concentrations of viral RNA during persistence were detected in the caecum and colon, suggesting that viral persistence is maintained in these tissues. Possible adaptive changes arising during persistence in vivo appeared to accumulate in the minor capsid protein (VP2) and the viral polymerase (NS7), in contrast with adaptive mutations selected during cell-culture passages in RAW264.7 cells that appeared in the major capsid protein (VP1) and non-structural protein NS4. This system provides an attractive model that can be readily used to identify viral sequences that contribute to persistence in an immunocompetent host and to more acute infection in an immunocompromised host, providing new insights into the biology of norovirus infections.
doi:10.1099/vir.0.042176-0
PMCID: PMC3541523  PMID: 22495235
12.  Retraction 
The Journal of General Virology  2010;91(Pt 10):2658.
doi:10.1099/vir.0.84530-0
PMCID: PMC3378246  PMID: 20847365
13.  Establishment of productively infected walleye dermal sarcoma explant cells 
The Journal of General Virology  2007;88(Pt 9):2583-2589.
Walleye dermal sarcoma virus (WDSV) is a complex retrovirus associated with dermal sarcomas in walleye fish. Virus expression is tightly regulated and limited to accessory gene transcripts throughout tumour development. During tumour regression, this regulation is lost and the replication of virus is greatly enhanced. Cultured walleye fibroblasts infected in vitro do not produce significant quantities of infectious virus. Tissue culture cells established by explantation of tumour cells were found to harbour WDSV provirus and to express accessory and structural proteins. The sequence of the provirus showed little variation from a previous WDSV isolate. Retroviral particles were isolated from supernatants from these cells and were able to transfer infection to uninfected walleye fibroblasts. In addition to the virus present in supernatants, much of the virus was cell associated and liberated only by sonication. This virus was found at internal cellular membranes, including mitochondria, and was infectious.
doi:10.1099/vir.0.82967-0
PMCID: PMC3364294  PMID: 17698670
14.  The origin and phylogeography of dog rabies virus 
The Journal of General Virology  2008;89(Pt 11):2673-2681.
Rabies is a progressively fatal and incurable viral encephalitis caused by a lyssavirus infection. Almost all of the 55 000 annual rabies deaths in humans result from infection with dog rabies viruses (RABV). Despite the importance of rabies for human health, little is known about the spread of RABV in dog populations, and patterns of biodiversity have only been studied in limited geographical space. To address these questions on a global scale, we sequenced 62 new isolates and performed an extensive comparative analysis of RABV gene sequence data, representing 192 isolates sampled from 55 countries. From this, we identified six clades of RABV in non-flying mammals, each of which has a distinct geographical distribution, most likely reflecting major physical barriers to gene flow. Indeed, a detailed analysis of phylogeographic structure revealed only limited viral movement among geographical localities. Using Bayesian coalescent methods we also reveal that the sampled lineages of canid RABV derive from a common ancestor that originated within the past 1500 years. Additionally, we found no evidence for either positive selection or widespread population bottlenecks during the global expansion of canid RABV. Overall, our study reveals that the stochastic processes of genetic drift and population subdivision are the most important factors shaping the global phylogeography of canid RABV.
doi:10.1099/vir.0.2008/003913-0
PMCID: PMC3326349  PMID: 18931062
15.  Comparative analysis reveals frequent recombination in the parvoviruses 
The Journal of General Virology  2007;88(Pt 12):3294-3301.
Parvoviruses are small single-stranded DNA viruses that are ubiquitous in nature. Infections with both autonomous and helper-virus dependent parvoviruses are common in both human and animal populations, and many animals are host to a number of different parvoviral species. Despite the epidemiological importance of parvoviruses, the presence and role of genome recombination within or among parvoviral species has not been well characterized. Here we show that natural recombination may be widespread in these viruses. Different genome regions of both porcine parvoviruses and Aleutian mink disease viruses have conflicting phylogenetic histories, providing evidence for recombination within each of these two species. Further, the rodent parvoviruses show complex evolutionary histories for separate genomic regions, suggesting recombination at the interspecies level.
doi:10.1099/vir.0.83255-0
PMCID: PMC3326350  PMID: 18024898
16.  Adaptation of two flaviviruses results in differences in genetic heterogeneity and virus adaptability 
The Journal of general virology  2007;88(Pt 9):2398-2406.
West Nile virus (WNV) is a mosquito-borne flavivirus that was first introduced into the USA in the New York City area in 1999. Since its introduction, WNV has steadily increased both its host and geographical ranges. Outbreaks of the closely related flavivirus, St. Louis encephalitis virus (SLEV), occur in the USA periodically, but levels of activity and host range are more restricted than those of WNV. Understanding the selective pressures that drive arbovirus adaptation and evolution in their disparate mosquito and avian hosts is crucial to predicting their ability to persist and re-emerge. Here, we evaluated the in vivo phenotypes of mosquito cell-adapted WNV and SLEV. Results indicated that in vitro adaptations did not translate to in vivo adaptations for either virus, yet SLEV displayed attenuated growth in both mosquitoes and chickens, while WNV generally did not. In vitro growth analyses also indicated that WNV adaptations could be generalized to cell cultures derived from other mosquito species, while SLEV could not. Analysis of genetic diversity for passaged SLEV revealed a highly homogeneous population that differed significantly from previous results of high levels of diversity in WNV. We hypothesize that this difference in genetic diversity is directly related to the viruses’ success in new and changing environments in the laboratory and that differences in a viruses’ ability to produce and maintain heterogeneous populations in nature may in some instances explain the variable levels of success seen among arboviruses.
doi:10.1099/vir.0.83061-0
PMCID: PMC3249635  PMID: 17698648
17.  Characterization of mosquito-adapted West Nile virus 
The Journal of general virology  2008;89(Pt 7):1633-1642.
West Nile virus (WNV), a mosquito-borne flavivirus, has significantly expanded its geographical and host range since its 1999 introduction into North America. The underlying mechanisms of evolution of WNV and other arboviruses are still poorly understood. Studies evaluating virus adaptation and fitness in relevant in vivo systems are largely lacking. In order to evaluate the capacity for host-specific adaptation and the genetic correlates of adaptation in vivo, this study measured phenotypic and genotypic changes in WNV resulting from passage in Culex pipiens mosquitoes. An increase in replicative ability of WNV in C. pipiens was attained for the two lineages of WNV tested. This adaptation for replication in mosquitoes did not result in a replicative cost in chickens, but did decrease cell-to-cell spread of virus in vertebrate cell culture. Genetic analyses of one mosquito-adapted lineage revealed a total of nine consensus nucleotide substitutions with no accumulation of a significant mutant spectrum. These results differed significantly from previous in vitro studies. When St Louis encephalitis virus (SLEV), a closely related flavivirus, was passaged in C. pipiens, moderately attenuated growth in C. pipiens was observed for two lineages tested. These results suggest that significant differences in the capacity for mosquito adaptation may exist between WNV and SLEV, and demonstrate that further comparative studies in relevant in vivo systems will help elucidate the still largely unknown mechanisms of arboviral adaptation in ecologically relevant hosts.
doi:10.1099/vir.0.2008/000893-0
PMCID: PMC3249650  PMID: 18559933
18.  Role of the mutant spectrum in adaptation and replication of West Nile virus 
The Journal of general virology  2007;88(Pt 3):865-874.
West Nile virus (WNV) has successfully spread throughout the USA, Canada, Mexico, the Caribbean and parts of Central and South America since its 1999 introduction into North America. Despite infecting a broad range of both mosquito and avian species, the virus remains highly genetically conserved. This lack of evolutionary change over space and time is common with many arboviruses and is frequently attributed to the adaptive constraints resulting from the virus cycling between vertebrate hosts and invertebrate vectors. WNV, like most RNA viruses studied thus far, has been shown in nature to exist as a highly genetically diverse population of genotypes. Few studies have directly evaluated the role of these mutant spectra in viral fitness and adaptation. Using clonal analysis and reverse genetics experiments, this study evaluated genotype diversity and the importance of consensus change in producing the adaptive phenotype of WNV following sequential mosquito cell passage. The results indicated that increases in the replicative ability of WNV in mosquito cells correlate with increases in the size of the mutant spectrum, and that consensus change is not solely responsible for alterations in viral fitness and adaptation of WNV. These data provide evidence of the importance of quasispecies dynamics in the adaptation of a flavivirus to new and changing environments and hosts, with little evidence of significant genetic change.
doi:10.1099/vir.0.82606-0
PMCID: PMC3249657  PMID: 17325359
19.  OTK18, a zinc finger protein, regulates human immunodeficiency virus type I long terminal repeat through two distinct regulatory regions 
The Journal of general virology  2007;88(Pt 1):236-241.
Summary
We have previously shown that OTK18, a human immunodeficiency virus (HIV)-inducible zinc finger protein, reduces progeny virion production in infected human macrophages. OTK18 antiviral activity is mediated through suppression of Tat-induced HIV-1 viral long terminal repeat (LTR) promoter activity. Through the use of LTR scanning mutant vectors, we have defined the specific regions responsible for OTK18-mediated LTR suppression. Two different LTR regions were identified as potential OTK18 binding sites by an enhanced DNA-transcription factor ELISA system; the negative regulatory element (NRE) at -255 / −238 and Ets binding site (EBS) at −150 / −139 in the LTR. In addition, deletion of the EBS in the LTR blocked OTK18-mediated LTR suppression. These data indicate that OTK18 suppresses LTR activity through two distinct regulatory elements. Spontaneous mutations in these regions might enable HIV-1 to escape from OTK18-antiretroviral activity in human macrophages.
doi:10.1099/vir.0.82066-0
PMCID: PMC3229096  PMID: 17170456
21.  Development and Characterization of Promoterless Helper RNAs for Production of Alphavirus Replicon Particles 
The Journal of general virology  2010;91(Pt 7):1723-1727.
SUMMARY
Alphavirus-based replicon systems are frequently used as preclinical vectors and as antigen discovery tools, and have recently been assessed in clinical vaccine trials. Typically, alphavirus replicon RNAs are delivered within virus-like replicon particles (VRP) that are produced following transfection of replicon RNA and two helper RNAs into permissive cells in vitro. The nonstructural proteins expressed from the replicon RNA amplify the replicon RNA in cis and the helper RNAs in trans, the latter providing the viral structural proteins necessary to package the replicon RNA into VRP. Current helper RNA designs incorporate the alphavirus 26S promoter to direct the transcription of high levels of structural-gene mRNAs. We demonstrate here that the 26S promoter is not required on helper RNAs to produce VRP and propose that such promoterless helper RNAs, by design, reduce the probability of generating replication-competent virus that may otherwise result from RNA recombination.
doi:10.1099/vir.0.020081-0
PMCID: PMC2888770  PMID: 20181749
22.  Development and characterization of promoterless helper RNAs for the production of alphavirus replicon particle 
The Journal of General Virology  2010;91(Pt 7):1723-1727.
Alphavirus-based replicon systems are frequently used as preclinical vectors and as antigen discovery tools, and they have recently been assessed in clinical vaccine trials. Typically, alphavirus replicon RNAs are delivered within virus-like replicon particles (VRP) that are produced following transfection of replicon RNA and two helper RNAs into permissive cells in vitro. The non-structural proteins expressed from the replicon RNA amplify the replicon RNA in cis and the helper RNAs in trans, the latter providing the viral structural proteins necessary to package the replicon RNA into VRP. Current helper RNA designs incorporate the alphavirus 26S promoter to direct the transcription of high levels of structural gene mRNAs. We demonstrate here that the 26S promoter is not required on helper RNAs to produce VRP and propose that such promoterless helper RNAs, by design, reduce the probability of generating replication-competent virus that may otherwise result from RNA recombination.
doi:10.1099/vir.0.020081-0
PMCID: PMC2888770  PMID: 20181749
23.  Characterization of the Ebola Virus Nucleoprotein-RNA Complex 
The Journal of general virology  2010;91(Pt 6):1478-1483.
Summary
When Ebola virus nucleoprotein (NP) is expressed in mammalian cells, it assembles into helical structures. Here, the recombinant NP helix purified from cells expressing NP was characterized biochemically and morphologically. We found that the recombinant NP helix is associated with non-viral RNA which is not protected from RNase digestion and that the morphology of the helix changes depending on the environmental salt concentration. The N-terminal 450 amino acid residues of NP are sufficient for these properties. However, digestion of the NP-associated RNA eliminates the plasticity of the helix, suggesting that this RNA is an essential structural component of the helix, binding to individual NP molecules via the N-terminal 450 amino acids. These findings enhance our knowledge of Ebola virus assembly and understanding of the Ebola virus life cycle.
doi:10.1099/vir.0.019794-0
PMCID: PMC2878588  PMID: 20164259
24.  A mutation in H5 hemagglutinin that conferred human receptor recognition is not stably maintained during duck passage 
The Journal of general virology  2010;91(Pt 6):1461-1463.
Summary
A/Hong Kong/213/97 (HK213; H5N1), isolated from a human, binds to both avian- and human-type receptors due to an HA mutation likely acquired during adaptation to humans. Duck passage of this virus conferred lethality in ducks. Sequence analyses of the duck-passaged virus revealed that its HA gene reverted back to one recognizing only avian-type receptors, and consequently it bound human tissue to a lesser extent. This finding suggests that viruses with human-type receptor specificity are unlikely to be maintained in waterfowl, unlike those with the human-type PB2 mutation, such as H5N1 viruses of the Qinghai lineage.
doi:10.1099/vir.0.018572-0
PMCID: PMC2881941  PMID: 20130132
25.  Characterization of the Ebola virus nucleoprotein–RNA complex 
The Journal of General Virology  2010;91(Pt 6):1478-1483.
When Ebola virus nucleoprotein (NP) is expressed in mammalian cells, it assembles into helical structures. Here, the recombinant NP helix purified from cells expressing NP was characterized biochemically and morphologically. We found that the recombinant NP helix is associated with non-viral RNA, which is not protected from RNase digestion and that the morphology of the helix changes depending on the environmental salt concentration. The N-terminal 450 aa residues of NP are sufficient for these properties. However, digestion of the NP-associated RNA eliminates the plasticity of the helix, suggesting that this RNA is an essential structural component of the helix, binding to individual NP molecules via the N-terminal 450 aa. These findings enhance our knowledge of Ebola virus assembly and understanding of the Ebola virus life cycle.
doi:10.1099/vir.0.019794-0
PMCID: PMC2878588  PMID: 20164259

Results 1-25 (126)