bluuebunny/crossref_metadata_2025_split huggingface dataset:
Analysis of the complete genome sequence of black queen-cell virus, a picorna-like virus of honey bees
Authors: Leat Neil , Ball Brenda , Govan Vandana , Davison Sean 
Published: August 2015

A virus with picorna-like biophysical properties was isolated from South African honey bees. On the basis of serology, it was identified as an isolate of black queen-cell virus (BQCV). Nucleotide sequence analysis revealed an 8550 nt polyadenylated genome containing two large ORFs. The 5′-proximal ORF (ORF 1) represented 4968 nt while the 3′-proximal ORF (ORF 2) represented 2562 nt. The ORFs were separated by a 208 nt intergenic region and were flanked by a 657 nt 5′-untranslated region and a 155 nt 3′-untranslated region. Deduced amino acid sequences for ORF 1 and ORF 2 were most similar to the non-structural and structural proteins, respectively, of Drosophila C virus (DCV), Rhopalosiphum padi virus (RhPV), Himetobi P virus (HiPV) and Plautia stali intestine virus (PSIV). It is proposed that BQCV belongs to the group of picorna-like, insect-infecting RNA viruses constituted by DCV, RhPV, HiPV and PSIV.
Open reading frame 2 of porcine circovirus type 2 encodes a major capsid protein
Authors: Nawagitgul Porntippa , Morozov Igor , Bolin Steven R. , Harms Perry A. , Sorden Steven D. , Paul Prem S. 
Published: August 2015

Porcine circovirus 2 (PCV2), a single-stranded DNA virus associated with post-weaning multisystemic wasting syndrome of swine, has two potential open reading frames, ORF1 and ORF2, greater than 600 nucleotides in length. ORF1 is predicted to encode a replication-associated protein (Rep) essential for replication of viral DNA, while ORF2 contains a conserved basic amino acid sequence at the N terminus resembling that of the major structural protein of chicken anaemia virus. Thus far, the structural protein(s) of PCV2 have not been identified. In this study, a viral structural protein of 30 kDa was identified in purified PCV2 particles. ORF2 of PCV2 was cloned into a baculovirus expression vector and the gene product was expressed in insect cells. The expressed ORF2 gene product had a molecular mass of 30 kDa, similar to that detected in purified virus particles. The recombinant ORF2 protein self-assembled to form capsid-like particles when viewed by electron microscopy. Antibodies against the ORF2 protein were detected in samples of sera obtained from pigs as early as 3 weeks after experimental infection with PCV2. These results show that the major structural protein of PCV2 is encoded by ORF2 and has a molecular mass of 30 kDa.
Nucleotide sequence, genome organization and phylogenetic analysis of pineapple mealybug wilt-associated virus-2
Authors: Melzer M. J. , Karasev A. V. , Sether D. M. , Hu J. S. 
Published: August 2015

The genome of pineapple mealybug wilt-associated closterovirus-2 (PMWaV-2) was cloned from double-stranded RNA isolated from diseased pineapple and its sequence determined. The 3′-terminal 14861 nt of the single-stranded RNA genome contains ten open reading frames (ORFs) which, from 5′ to 3′, potentially encode a >204 kDa polyprotein containing papain-like protease, methyltransferase and helicase domains (ORF1a), a 65 kDa RNA-dependent RNA polymerase (ORF1b), a 5 kDa hydrophobic protein (ORF2), a 59 kDa heat shock protein 70 homologue (ORF3), a 46 kDa protein (ORF4), a 34 kDa coat protein (ORF5), a 56 kDa diverged coat protein (ORF6), a 20 kDa protein (ORF7), a 22 kDa protein (ORF8) and a 6 kDa protein (ORF9). A 132 nt untranslated region was present at the 3′ terminus of the genome. This genome organization is typical of the monopartite closteroviruses, including the putative +1 ribosomal frameshift allowing expression of ORF1b. Phylogenetic analysis revealed that within the family Closteroviridae the mealybug-transmitted PMWaV-2 is more closely related to other mealybug-transmitted members than to those which are transmitted by aphids or whiteflies. Within this group, PMWaV-2 shares the greatest sequence identity with grapevine leafroll-associated virus-3, another mealybug-transmitted closterovirus.
Construction and characterization of recombinant porcine adenovirus serotype 5 expressing the transmissible gastroenteritis virus spike gene
Authors: Tuboly Tamás , Nagy Éva 
Published: August 2015

Five recombinant porcine adenoviruses of serotype 5 (PAdV-5) carrying the full-length or the 5′ 2·2 kb half of the transmissible gastroenteritis virus (TGEV) spike (S) gene were generated by homologous recombination in E. coli strain BJ5183 cells and subsequent transfection of swine testicle cells. The foreign genes were inserted into the E3 region of PAdV-5. One recombinant virus had no deletion in the E3 region, whereas a 1·2 kb fragment was removed from the E3 region in the remainder of the recombinant viruses. One stable construct with a 4·4 kb insertion had a genome size of 109·6% of the wild-type genome, the largest reported for any recombinant adenovirus. Only those viruses that carried the S gene in the left to right orientation expressed the S gene. Three recombinant viruses were tested by oral immunization of pigs and both antibody response and virus shedding were monitored. None of the pigs showed clinical signs and the virus was recovered from rectal swabs until 6–7 days post-infection. Viruses expressing the S gene induced TGEV- and PAdV-5-specific virus-neutralizing antibodies. Moreover, TGEV-specific secretory IgA was detected in the small intestine and in the lungs of the immunized animals.
The VP1-unique region of parvovirus B19: amino acid variability and antigenic stability
Authors: Dorsch Simone , Kaufmann Bärbel , Schaible Uwe , Prohaska Elke , Wolf Hans , Modrow Susanne 
Published: August 2015

The unique region of structural protein VP1 of parvovirus B19 (erythrovirus B19) is important for eliciting neutralizing antibodies that are responsible for eliminating the virus from the peripheral blood and for inducing lifelong immunity. Neutralizing human MAbs bind a conformationally defined epitope spanning VP1 residues 30–42. The DNA sequence encoding the VP1-unique region was determined in parvovirus B19 isolated from peripheral blood and amniotic fluid of nine acutely infected pregnant women, five arthritis patients and two chronically infected children. The amino acid sequences of the VP1-unique region exhibited higher variability in comparison with other B19-specific proteins. To analyse the influence of amino acid variations on antibody binding and protein conformation, two variants of the VP1-unique region were selected and expressed in E. coli as intein-fusion proteins. The selected variants displayed a number of amino acid exchanges in the VP1-unique region and had mutations in the determined epitope and adjacent regions. After purification via affinity chromatography, the dissociation constants K
D of VP1-specific human MAbs interacting with the variant antigens and a viral prototype of the VP1-unique region were determined with a quartz crystal microbalance biosensor. A value of 5·4×10−8 M was determined for the prototype isolate pJB; the affinity constants for the variant VP1-unique regions were similar. Comparable values were obtained for interaction of antibodies with non-infectious VP1/VP2 capsids produced by recombinant baculovirus and with B19 virions from amniotic fluid. It is concluded that the conformation of the epitope is unaffected by mutations or the environment of the VP1-unique region in virus capsids.
Analysis of the capsid processing strategy of Thosea asigna virus using baculovirus expression of virus-like particles
Authors: Pringle Fiona M. , Kalmakoff James , Ward Vernon K. 
Published: August 2015

Thosea asignavirus (TaV), a putative member of the genusBetatetravirusof the familyTetraviridae, is predicted to have a novel capsid expression strategy compared with other characterized tetraviruses. The capsid precursor protein is cleaved twice to generate three proteins. Two of the proteins, L (58·3 kDa) and S (6·8 kDa), are incorporated into the TaV virion. The third, non-structural protein, produced from the N terminus of the precursor protein, is up to 17 kDa in size and is of unknown function. The TaV capsid precursor protein sequence without the 17 kDa N-terminal region was modelled against the solved structure fromNudaureliaω virus (NωV) using SwissModel. The TaV model was very similar to the solved structure determined for subunit A of NωV and had features that are conserved between tetraviruses and nodaviruses, including the positioning of the cleavage site between the L and S capsid proteins. The production of virus-like particles (VLPs) using the baculovirus expression system was used to analyse the capsid processing strategy employed by TaV. VLPs were formed in both the presence and absence of the 17 kDa N-terminal region of the capsid precursor. VLPs were not formed when the L and S regions were expressed from separate promoters, indicating that cleavage between the L and S capsid proteins was an essential part of TaV capsid assembly. Expression of the TaV 17 kDa protein in bacteria did not produce intracellular tubules similar to those formed by bacterial expression of the p17 protein fromHelicoverpa armigerastunt virus.
Complete nucleotide sequence and host range of South African cassava mosaic virus: further evidence for recombination amongst begomoviruses
Authors: Berrie L. C. , Rybicki E. P. , Rey M. E. C. 
Published: August 2015

Complete nucleotide sequences of the DNA-A (2800 nt) and DNA-B (2760 nt) components of a novel cassava-infecting begomovirus, South African cassava mosaic virus (SACMV), were determined and compared with various New World and Old World begomoviruses. SACMV is most closely related to East African cassava mosaic virus (EACMV) in both its DNA-A (85% with EACMV-MH and -MK) and -B (90% with EACMV-UG2-Mld and EACMV-UG3-Svr) components; however, percentage sequence similarities of less than 90% in the DNA-A component allowed SACMV to be considered a distinct virus. One significant recombination event spanning the entire AC4 open reading frame was identified; however, there was no evidence of recombination in the DNA-B component. Infectivity of the cloned SACMV genome was demonstrated by successful agroinoculation of cassava and three other plant species (Phaseolus vulgaris, Malva parviflora and Nicotiana benthamiana). This is the first description of successful infection of cassava with a geminivirus using Agrobacterium tumefaciens.
Molecular strategy for ‘serotyping’ of human enteroviruses
Authors: Caro Valérie , Guillot Sophie , Delpeyroux Francis , Crainic Radu 
Published: August 2015

To explore further the phylogenetic relationships between human enteroviruses and to develop new diagnostic approaches, we designed a pair of generic primers in order to study a 1452 bp genomic fragment (relative to the poliovirus Mahoney genome), including the 3′ end of the VP1-coding region, the 2A- and 2B-coding regions, and the 5′ moiety of the 2C-coding region. Fifty-nine of the 64 prototype strains and 45 field isolates of various origins, involving 21 serotypes and 6 strains untypeable by standard immunological techniques, were successfully amplified with these primers. By determining the nucleotide sequence of the genomic fragment encoding the C-terminal third of the VP1 capsid protein we developed a molecular typing method based on RT–PCR and sequencing. If field isolate sequences were compared to human enterovirus VP1 sequences available in databases, nucleotide identity score was, in each case, highest with the homotypic prototype (74.8 to 89.4%). Phylogenetic trees were generated from alignments of partial VP1 sequences with several phylogeny algorithms. In all cases, the new classification of enteroviruses into five identified species was confirmed and strains of the same serotype were always monophyletic. Analysis of the results confirmed that the 3′ third of the VP1-coding sequence contains serotype-specific information and can be used as the basis of an effective and rapid molecular typing method. Furthermore, the amplification of such a long genomic fragment, including non-structural regions, is straightforward and could be used to investigate genome variability and to identify recombination breakpoints or specific attributes of pathogenicity.
E5 transforming proteins of papillomaviruses do not disturb the activity of the vacuolar H+-ATPase
Authors: Ashby Andrew D. M. , Meagher Liam , Campo M. Saveria , Finbow Malcolm E. 
Published: August 2015

Papillomaviruses contain a gene, E5, that encodes a short hydrophobic polypeptide that has transforming activity. E5 proteins bind to the 16 kDa subunit c (proteolipid) of the eukaryotic vacuolar H+-ATPase (V-ATPase) and this binding is thought to disturb the V-ATPase and to be part of transformation. This link has been examined in the yeastSaccharomyces cerevisiae. The E5 proteins from human papillomavirus (HPV) type 16, bovine papillomavirus (BPV) type 1, BPV-4 E5 and various mutants of E5 and the p12′ polypeptide from human T-lymphotropic virus (HTLV) type I all bound to theS. cerevisiaesubunit c (Vma3p) and could be found in vacuolar membranes. However, none affected the activity of the V-ATPase. In contrast, a dominant-negative mutant of Vma3p (E137G) inactivated the enzyme and gave the characteristic VMA phenotype. A hybrid V-ATPase containing a subunit c from Norway lobster also showed no disruption. Sedimentation showed that HPV-16 E5 was not part of the active V-ATPase. It is concluded that the binding of E5 and E5-related proteins to subunit c does not affect V-ATPase activity or function and it is proposed that the binding may be due to a chaperone function of subunit c.
Evaluation of a prototype sub-unit vaccine against equine arteritis virus comprising the entire ectodomain of the virus large envelope glycoprotein (GL): induction of virus-neutralizing antibody and assessment of protection in ponies
Authors: Castillo-Olivares J. , de Vries A. A. F. , Raamsman M. J. B. , Rottier P. J. M. , Lakhani K. , Westcott D. , Tearle J. P. , Wood J. L. N. , Mumford J. A. , Hannant D. , Davis-Poynter N. J. 
Published: August 2015

An Escherichia coli-expressed recombinant protein (6hisGLecto) comprising the entire ectodomain (aa 18–122) of equine arteritis virus (EAV) glycoprotein GL, the immunodominant viral antigen, induced higher neutralizing antibody titres than other GL-derived polypeptides when compared in an immunization study in ponies. The potential of the recombinant GL ectodomain to act as a sub-unit vaccine against EAV was evaluated further in three groups of four ponies vaccinated with doses of 35, 70 or 140 μg of protein. All vaccinated animals developed a virus-neutralizing antibody (VNAb) response with peak titres 1–2 weeks after the administration of a booster on week 5 (VNAb titres of 1·8–3·1), 13 (VNAb titres of 1·4–2·9) or 53 (VNAb titres of 1·2–2·3). Vaccinated and unvaccinated control ponies were infected with EAV at different times post-vaccination to obtain information about the degree of protection relative to the levels of pre-challenge VNAb. Vaccination conferred varying levels of protection, as indicated by reduced or absent pyrexia, viraemia and virus excretion from the nasopharynx. The degree of protection correlated well with the levels of pre-challenge VNAb and, in particular, with levels of virus excretion. These results provide the first evidence that a sub-unit vaccine protects horses against EAV. The use of the sub-unit vaccine in combination with a differential diagnostic test based on other EAV antigens would enable serological discrimination between naturally infected and vaccinated equines.
The complete sequence of the Cydia pomonella granulovirus genome
Authors: Luque Teresa , Finch Ruth , Crook Norman , O’Reilly David R. , Winstanley Doreen 
Published: August 2015

The nucleotide sequence of the DNA genome of Cydia pomonella granulovirus (CpGV) was determined and analysed. The genome is composed of 123500 bp and has a G+C content of 45·2%. It contains 143 ORFs of 150 nucleotides or more that show minimal overlap. One-hundred-and-eighteen (82·5%) of these putative genes are homologous to genes previously identified in other baculoviruses. Among them, 73 are homologous to genes of Autographa californica nucleopolyhedrovirus (AcMNPV), whereas 108 and 98 are homologous to genes of Xestia c-nigrum GV (XcGV) and Plutella xylostella GV (PxGV), respectively. These homologues show on average 37·4% overall amino acid sequence identity to those from AcMNPV and 45% to those from XcGV and PxGV. The CpGV gene content was compared to that of other baculoviruses. Several genes reported to have major roles in baculovirus biology were not found in the CpGV genome, such as gp64, the major budded virus glycoprotein gene in some nucleopolyhedroviruses, and lef-7, involved in DNA replication. However, the CpGV genome encodes the large and small subunits of ribonucleotide reductase, three inhibitor of apoptosis (iap) homologues and two protein tyrosine phosphatases. The CpGV, PxGV and XcGV genomes present a noticeably high level of conservation of gene order and orientation. A striking feature of the CpGV genome is the absence of typical homologous repeat sequences. However, it contains one major repeat region and 13 copies of a single 73–77 bp imperfect palindrome.
Cloning and sequence analysis of an infectious clone of Citrus yellow mosaic virus that can infect sweet orange via Agrobacterium-mediated inoculation
Authors: Huang Qi , Hartung John S. 
Published: August 2015

Citrus yellow mosaic virus(CYMV), a member of the familyCaulimoviridae, genusBadnavirus, causes citrus mosaic disease, a disease that occurs commonly in India. The CYMV genome has been cloned and its complete nucleotide sequence determined. Its DNA genome is 7559 bp in length and contains six putative open reading frames (ORFs), all on the plus-strand of the genome and each capable of encoding proteins with a molecular mass of greater than 10 kDa. ORF 3, the largest ORF, encodes a putative polyprotein for functions involved in virus movement, assembly and replication. The other ORFs encode proteins whose exact functions are not completely understood. The genome also contains a plant tRNAmet-binding site, which may serve as a primer for minus-strand DNA synthesis, in its intergenic region. Phylogenetic analysis of the badnaviruses revealed that CYMV is most closely related toCacao swollen shoot virus. It was demonstrated that a construct containing 1·4 copies of the cloned CYMV genome could infect sweet orange viaAgrobacterium-mediated inoculation.
Increased replication of respiratory syncytial virus (RSV) in pulmonary infiltrates is associated with enhanced histopathological disease in bonnet monkeys (Macaca radiata) pre-immunized with a formalin-inactivated RSV vaccine
Authors: Ponnuraj Esther M. , Hayward Anthony R. , Raj Anthony , Wilson Harry , Simoes Eric A. F. 
Published: August 2015

The pathology of respiratory syncytial virus (RSV) disease in bonnet monkeys parallels findings with human RSV disease. RSV-infected animals pre-immunized with a formalin-inactivated (FI) RSV vaccine develop inflammation in peribronchiolar, perivascular, interstitial and intra-alveolar sites with lung inflammation scores significantly higher than animals with a primary RSV infection and those pre-immunized with an FI-Vero cell control vaccine (P=0·05). Animals previously infected and re-exposed to RSV had significantly lower alveolar, interstitial and total lung inflammation scores than in primary infection (P=0·05). Immunization with two intra-muscular doses of 0·5 ml of the FI-RSV vaccine administered 21 days apart resulted in little serum-neutralizing and ELISA antibody, low levels of secretory IgA and a low lymphocyte proliferative response that was significantly lower than the response observed in animals that were previously infected with live RSV. Higher RSV virus titres were detected in the lungs and lung lavage fluid of monkeys immunized with the FI-RSV vaccine than in those with a primary infection (P=0·001). RSV was detected byin situhybridization in pulmonary inflammatory infiltrates, where the single most abundant infiltrating cellular species was macrophages, so it may be these cells that support the enhanced virus replication that contributes to the enhanced pulmonary pathology of FI-RSV immunization.
Varicella-zoster virus gH:gL contains a structure reactive with the anti-human gamma chain of IgG near the glycosylation site
Authors: Yokoyama Tomonori , Ayabe Satoko , Miyagi Huminori , Sugano Toru , Otsu Akira , Sato Hitoshi , Kageyama Seiji , Fujii Takao , Shiraki Kimiyasu 
Published: August 2015

Varicella-zoster virus (VZV) glycoproteins were purified from infected cells using monoclonal antibodies and gH:gL was found to react with antibodies to the γ chain of human IgG (h-IgG), whereas gE:gI and gB did not. When gH:gL was captured by concanavalin A, it lost reactivity with the anti-h-IgG γ chain (anti-h-γ-IgG). gH:gL reacted with anti-h-γ-IgG in an ELISA assay and gave a K
d value of 2·16×10−7 M in a BIAcore assay. The K
d value of the human monoclonal antibody to gH (TI-57) used for the purification of gH:gL was 4·45×10−10 M. Virus pretreated with anti-h-IgG was five times more resistant to neutralization with TI-57. Although the nature of the binding was not clear, gH:gL bound to anti-h-γ-IgG. If this interaction results from immunological similarity between gH:gL and h-IgG, it may cause immune evasion in the pathogenesis of VZV infection.
Identification and characterization of a new intron in Borna disease virus
Authors: Cubitt Beatrice , Ly Calvin , de la Torre Juan Carlos 
Published: August 2015

Borna disease virus (BDV) has a non-segmented, negative-strand (NNS) RNA genome. In contrast to all other known NNS RNA animal viruses, BDV replication and transcription occur in the nucleus of infected cells. Moreover, BDV uses RNA splicing for the regulation of its genome expression. Two introns (I and II), both present in two viral primary transcripts of 2·5 and 7·2 kb, have been reported in BDV. Here, evidence is provided of a new BDV intron, intron III, generated by alternative 3′ splice-site choice. Intron III-spliced mRNAs were detected at early times post-infection and found to be present in cells from different types and species. Intron III-spliced mRNAs have coding capability for two new viral proteins with predicted molecular masses of 8·4 and 165 (p165) kDa. p165 is a deleted form of the BDV L polymerase, containing three RGD motifs and a signal peptide signal that could target it into the secretory pathway. These findings underscore the proteomic complexity exhibited by BDV.