Virus research.
Publisher:
Elsevier Science,
Frequency: 16 no. a year, 1999-
Country: Netherlands
Language: English
Start Year:1984 -
ISSN:
0168-1702 (Print)
1872-7492 (Electronic)
0168-1702 (Linking)
1872-7492 (Electronic)
0168-1702 (Linking)
Impact Factor
5
2022
| NLM ID: | 8410979 |
| (DNLM): | V08043000(s) |
| (OCoLC): | 10694473 |
| Coden: | VIREDF |
| Classification: | W1 VI836D |
The effect of siRNA treatment on experimental equine herpesvirus type 1 (EHV-1) infection in horses. Available vaccines fail to induce lasting and protective immunity to equine herpesvirus 1 (EHV-1) associated diseases. RNA interference is a novel approach showing promise for therapeutic use in outbreak situations. This study examined the effect of small interfering RNA (siRNA) on clinical signs as well as the presence of live virus and viral DNA in nasal secretions and peripheral blood mononuclear cells (PBMCs) in horses experimentally infected with EHV-1. siRNA targeting two EHV-1 genes (glycoprotein B and the origin binding protein) was administered 12h before and 12h after intranasal infe...
Small interfering RNA targeting bovine papillomavirus type 1 E2 induces apoptosis in equine sarcoid transformed fibroblasts. Equine sarcoids are skin tumours of horses caused by infection with BPV-1 or 2. Maintenance and replication of the viral genome depend upon the viral proteins E1 and E2. We examined the effects of an E2 specific siRNA on E2 and E1 viral gene expression, viral load and cell growth in BPV-1 transformed sarcoid-derived cells. Transfection with E2-siRNA caused a reduction in E2 and E1 mRNA expression as well as viral load, growth inhibition and decreased anchorage-independent growth. siRNA treated cells showed significantly higher apoptosis rates than control cells. Thus sequence specific targetin...
Aggregation-associated loss of antigenicity observed for denatured virion protein 1 of Equine rhinitis A virus in an enzyme-linked immunosorbent assay. Equine rhinitis A virus (ERAV) is a picornavirus which causes an acute respiratory infection in horses worldwide, and virus neutralization (VN) has been the standard method for the detection of ERAV antibody in horse serum. Previous studies have identified recombinant virion protein VP1 (rVP1) purified under native conditions to be of high potential for the development of a diagnostic ERAV enzyme-linked immunosorbent assay (ELISA). This study presents an optimized protocol for the expression and purification of native full-length rVP1. Furthermore, we demonstrate that, upon denaturation, rVP1 ...
Virion associated proteins of equine rhinitis B virus 1 (ERBV1): the non-structural protein 3C(pro) co-purifies with virions. Equine rhinitis B virus (ERBV), genus Erbovirus, is most closely related to the Cardiovirus genus in the family Picornaviridae. The structural proteins (VP1-4) of erboviruses are not well described, but are predicted by sequence to be 35, 29, 26 and 7 kDa. Methods for the purification of cardioviruses (polyethylene glycol, trypsin treatment) were used to characterise the structural proteins of ERBV1. Only one of the virus proteins detected was an expected molecular mass, and this 26 kDa protein was identified as VP3 by N-terminal amino acid sequencing. N-terminal sequencing of the 56 and a 29 ...
Truncation of cytoplasmic tail of EIAV Env increases the pathogenic necrosis. Equine Infectious Anemia Virus (EIAV), like other lentiviruses, has a transmembrane glycoprotein with an unusually long cytoplasmic tail (CT). Viral envelope (Env) proteins having CT truncations just downstream the putative membrane-spanning domain (PMSD) are assumed to exist among all wild-type budded virions, and also in some cell-adapted strains. To determine whether CT-truncated Env proteins can cause particularly deleterious effects on the Env expressing cells and/or their neighboring cells, plasmids encoding codon-optimized env gene including full-length (pE863) or CT-truncated (pE686* a...
Genetic characterization of equine influenza viruses isolated in Italy between 1999 and 2005. During local respiratory disease outbreaks, occurring in 2003 and 2004 in horse training stables within race-tracks in Rome, and on a stud horse farm in Bari in 2005, four strains of equine influenza (EI) virus were isolated. All outbreaks occurred in flu-vaccinated horses. Here, we are reporting the results of the genetic characterization of these isolates, together with that of another EI virus strain isolated in 1999 from a dead foal presenting pulmonary lesions. Alignment and phylogenetic analyses were carried out using the haemagglutinin amino acid sequences. The Rome and Bari isolates we...
Gag genetic heterogeneity of equine infectious anemia virus (EIAV) in naturally infected horses in Canada. Gag genetic heterogeneity of equine infectious anemia virus (EIAV) variants in naturally infected horses in Canada was studied since very limited information is available on the variability of EIAV Gag sequences in public database. A phylogenetic analysis based on 414nts of Gag gene sequences amplified by a nested polymerase chain reaction (PCR) revealed the distinct divergence of these variants compared to other published strains in a corresponding region. Significant predicted amino acid sequence variations were also identified in an immunorelevant region within this fragment which correspon...
Long terminal repeat sequences from virulent and attenuated equine infectious anemia virus demonstrate distinct promoter activities. In the early 1970s, the Chinese Equine Infectious Anemia Virus (EIAV) vaccine, EIAV(DLA), was developed through successive passages of a wild-type virulent virus (EIAV(L)) in donkeys in vivo and then in donkey macrophages in vitro. EIAV attenuation and cell tropism adaptation are associated with changes in both envelope and long terminal repeat (LTR). However, specific LTR changes during Chinese EIAV attenuation have not been demonstrated. In this study, we compared LTR sequences from both virulent and attenuated EIAV strains and documented the diversities of LTR sequence from in vivo and in v...
Live-attenuated recombinant equine herpesvirus type 1 (EHV-1) induces a neutralizing antibody response against West Nile virus (WNV). The immunogenicity in horses of a recombinant equine herpesvirus type 1 (EHV-1) vaccine expressing West Nile virus (WNV) prM and E proteins was studied. To construct the recombinant EHV-1, two-step en passant mutagenesis was employed for manipulation of a bacterial artificial chromosome (BAC) of vaccine strain RacH. Recombinant EHV-1 stably expressed the WNV prM and E proteins as demonstrated by indirect immunofluorescence and Western blotting. In addition, growth properties in vitro of the EHV-1/WNV recombinant were found to not be significantly different from those of the parental virus. To ...
Detection of bovine papillomavirus type 1 genomes and viral gene expression in equine inflammatory skin conditions. Papillomaviruses are normally strictly species-specific and even under experimental conditions do not usually infect any other host than the natural host. The only documented reports of natural papillomavirus cross-species infection are of BPV-1/BPV-2, which can infect horses and induce equine sarcoids. BPV DNA has not been detected in non-sarcoid equine tumours or equine papillomas, but its presence has been reported in some cases of equine dermatitis. In the present study, we show that equine inflammatory skin conditions harbour episomal circular double stranded BPV-1 genomes, with copy numb...
Molecular analysis of the proviral DNA of equine infectious anemia virus in mules in Greece. Molecular analysis of the regulatory and structurally important genetic segments of equine infectious anemia virus (EIAV) in mules is presented. We have previously reported clinicopathological and laboratory findings in mules infected with EIAV, both naturally and after experimental inoculation. In this study the fragment coding for integrase, gp90, tat and the fusion domain of gp45 of the proviral genome from these animals was sequenced and compared with one another and with that of EIAV strains already published in the literature. Significant variations were observed mainly in the sequences ...
Alternate circulation of recent equine-2 influenza viruses (H3N8) from two distinct lineages in the United States. Phylogenetic and antigenic analyses indicate that recent circulating equine-2 influenza viruses in the United States have been alternating between two genetic and antigenic distinct lineages since 1996. The evolution rates for these two lineages, the Kentucky and the Florida lineage, are very similar. For the earlier isolates in the Kentucky lineage, there are multiple and sequential nonsynonymous substitutions at antigenic sites B and D. However, there are no changes at any of these antigenic sites for KY98 and OK00. In the Florida lineage, except for NY99 with one amino acid substitution at ...
Generation and characterization of an EICP0 null mutant of equine herpesvirus 1. The EICP0 gene (gene 63) of equine herpesvirus 1 (EHV-1) encodes an early regulatory protein that is a promiscuous trans-activator of all classes of viral genes. Bacterial artificial chromosome (BAC) technology and RecE/T cloning were employed to delete the EICP0 gene from EHV-1 strain KyA. Polymerase chain reaction, Southern blot analysis, and DNA sequencing confirmed the deletion of the EICP0 gene and its replacement with a kanamycin resistance gene in mutant KyA. Transfection of rabbit kidney cells with the EICP0 mutant genome produced infectious virus, indicating that the EICP0 gene is not...
Sequence variants of bovine papillomavirus E5 detected in equine sarcoids. The equine sarcoid, one of the most common dermatological lesions in equids, is a benign, locally invasive dermal fibroblastic lesion. Previous studies have suggested an association with two bovine papilloma virus (BPV) types, BPV-1 and BPV-2. In the present study, we examined sarcoids from horses from two geographical areas, Switzerland and the UK, for the major transforming gene of BPV, E5. We detected BPV DNA for the E5 open reading frame and viral E5 RNA transcripts in most sarcoids. Sequence analysis of the E5 open reading frame of sarcoid-associated BPV detected several unique DNA sequen...
VP2 gene phylogenetic characterization of field isolates of African horsesickness virus serotype 7 circulating in South Africa during the time of the 1999 African horsesickness outbreak in the Western Cape. We present the first VP2-gene phylogenetic analysis of African horsesickness (AHS) viruses within a serotype. Thirteen AHSV 7 isolates were obtained from cases that occurred in South Africa during 1998-1999, and three were historical AHSV 7 isolates. The goals were to start a database of isolates of known location and time of isolation and to determine if we could identify the origin of an AHS outbreak in the surveillance area in the Western Cape. We prepared full-length cDNA copies of the VP2-genes of the isolates. Nucleic acid sequence data of a 786 bp region was used to characterize the gen...
Animal models of papillomavirus pathogenesis. Tumorigenesis due to papillomavirus (PV) infection was first demonstrated in rabbits and cattle early last century. Despite the evidence obtained in animals, the role of viruses in human cancer was dismissed as irrelevant. It took a paradigm shift in the late 1970s for some viruses to be recognised as 'tumour viruses' in humans, and in 1995, more than 60 years after Rous's first demonstration of CRPV oncogenicity, WHO officially declared that 'HPV-16 and HPV-18 are carcinogenic to humans'. Experimental studies with animal PVs have been a determining factor in this decision. Animal PVs have bee...
Detection of equine herpesvirus type 2 (EHV-2) in horses with keratoconjunctivitis. The prevalence of EHV-2 in 27 horses with keratoconjunctivitis and 21 clinically healthy horses of different ages and stocks were analyzed. We demonstrated that EHV-2 was present in 12 keratoconjunctivitis cases as shown by nested PCR on ocular swabs. This is statistically more often than in the control group, where only two ocular swabs were EHV-2 positive. Cocultivation was successful on peripheral blood leukocytes of healthy and diseased horses but not on swabs. We isolated ten EHV-2 strains from diseased and nine from control horses, whereas 16 isolates showed different restriction enzyme ...
Mutations occurring during serial passage of Japanese equine infectious anemia virus in primary horse macrophages. An attenuated equine infectious anemia virus (EIAV), named V26, was previously obtained after 50 passages of the Japanese virulent strain V70 in primary macrophage culture. To clarify the differences between both viruses, their full-length sequences were determined. There were higher mutations in S2 (6.15% amino acid difference) and LTR (10.7% nucleotide difference). The presumed initiation codon of the S2 gene was absent from the sequence of V26. There was a large insertion within the long-terminal repeat (LTR) U3 hypervariable region of V26. In addition, there were minor mutations in gag (1....
Bovine papillomaviral gene expression in equine sarcoid tumours. The sarcoid is a benign locally invasive dermal fibroblastic lesion, commonly affecting horses and donkeys. The aetiology of the equine sarcoid is equivocal. Bovine papillomaviral (BPV) DNA (type 1/2) is frequently demonstrable in equine sarcoid tumour biopsies. However, the exact role of the virus in the disease process and its contribution to the phenotypic differences in sarcoids is not known. It was sought to assess the transcriptional activity of BPV-1 found in sarcoid tissues. Of 20 tumours examined, 18 were positive for E2 expression and ten positive for L1 expression. Viral oncogenes E...
Virological and molecular biological investigations into equine herpes virus type 2 (EHV-2) experimental infections. Two 18-month-old naturally reared ponies were used to investigate the pathogenicity of EHV-2. After dexamethasone treatment, pony 1 was inoculated intranasally with EHV-2 strain T16, which has been isolated from a foal with keratoconjunctivitis superficialis and pony 2 was similarly inoculated with strain LK4 which was originally isolated from a horse with upper respiratory tract disease. Following virus inoculation, pyrexia was not detected in either pony but both developed conjunctivitis, lymphadenopathy, and coughing. EHV-2 was detected in nasal mucus samples up to day 12 post infection (p....
Sequence analysis of equine adenovirus 2 hexon and 23K proteinase genes indicates a phylogenetic origin distinct from equine adenovirus 1. We report the first nucleotide sequence data on equine adenovirus 2 (EAdV2) which corroborate on the molecular level that EAdV2 is distinct from equine adenovirus 1 (EAdV1). Based on sequence homology with Eadv1 the hexon gene of Eadv2 was identified. HindIII restriction fragments containing the hexon and eight other viral genes were cloned into the plasmid pUC19 and the nucleotide sequence of the hexon and the 23K proteinase genes completely determined. Amino acid (aa) comparison of sequence fragments with published adenovirus (AdV) proteins identified the genes for the E1B/19K, IVa2, DNA pol...
Varied prevalence of Borna disease virus infection in Arabic, thoroughbred and their cross-bred horses in Iran. Borna disease virus (BDV) naturally infects horses and sheep and induces progressive poliomeningoencephalomyelitis. Here, BDV recombinant proteins of the first open reading frame (ORF-I; coding for p40 nucleoprotein) and the second ORF-II (coding for p24 polymerase cofactor) were immunoblotted with plasma derived from 72 healthy (28 Arabic, 17 thoroughbred and 27 cross-bred) race horses at Tehran in Iran to detect anti-BDV antibodies. In addition, their peripheral blood mononuclear cells (PBMCs) were also examined for BDV RNA by a nested reverse transcriptase-polymerase chain reaction (RT-PCR)...
Ultrastructure of equine morbillivirus. The ultrastructure of the equine morbillivirus (EMV) which was implicated in the death of one human and fourteen horses in Queensland, Australia during September 1994 and a 36 year old man from Queensland in October 1995 is described. The ultrastructure of the virus and the intracellular virus-specific structures are characteristic for the family Paramyxoviridae. Cytoplasmic nucleocapsids were observed within the infected cells monolayers, endothelial cells (lung) of infected horses and the neurons within the brain of the 36 year old Queensland man. Aggregates of smaller nucleocapsid-like stru...
Comparison of the deduced matrix and fusion protein sequences of equine morbillivirus with cognate genes of the Paramyxoviridae. The nucleotide sequence of the matrix protein of equine morbillivirus (EMV) was determined to be 1062 nucleotides and coded for a deduced protein of M(r) 40148 having a net charge of + 19 at neutral pH. The matrix protein gene was separated from the P and F genes by intercistronic regions of 546 and 469 nucleotides, respectively. The nucleotide sequence which coded for the F protein was 1641 nucleotides and coded for a deduced protein of 546 amino acids having an M(r) of 60,447 and a charge + 4 at neutral pH. Partial sequence information was also determined for the P/V proteins. M, P and F pro...
Expression cloning and antigenic analysis of the nucleocapsid protein of equine arteritis virus. A series of recombinant fusion proteins derived from equine arteritis virus (EAV) open reading frame (ORF) 7 have been used to define the immunoreactive region of the viral nucleocapsid (N) protein. Reactivities of recombinant N fusion proteins with post-infection equine sera in immunoblots and ELISAs indicate that the major nucleocapsid protein epitope is located within amino acid residues 1-69. In ELISAs two recombinant nucleocapsid fusion proteins containing residues 1-69 (rN1-69) and 1-28 (rN1-28) discriminated between pre- and post-infection, and pre- and post-vaccination serum samples. A...
The use of African horse sickness virus NS3 protein, expressed in bacteria, as a marker to differentiate infected from vaccinated horses. Segment 10 of the double-stranded RNA (dsRNA) genome from African horse sickness virus serotype 4 (AHSV-4) was cloned and sequenced. The sequence of the coding region showed a total length of 667 bp. Nucleotide comparisons showed a 95% sequence similarity between serotypes 4 and 9, and 76% between serotypes 4 and 3. cDNA clones containing the coding region were cloned in the vector pET3xb and expressed in Escherichia coli. The NS3 gene product was synthesised at very high level as an insoluble fusion protein. The recombinant protein was used in a differential ELISA to distinguish horses that w...
Epitope mapping of cross-reactive monoclonal antibodies specific for the influenza A virus PA and PB2 polypeptides. Characterization of the epitopes recognized by 21 monoclonal antibodies (MAbs) specific for the influenza A virus PA (13 MAbs) and PB2 (8 MAbs) polypeptides (Bárcena et al. (1994) J. Virol. 68, 6900-6909) raised against denatured polypeptides produced in E. coli is described. MAbs were characterized by: (1) competitive binding ELISAs; (2) mapping of the protein regions that specify their binding sites; and (3) analyses of their ability to recognize the corresponding viral protein in a number of viral isolates. Five and three non-overlapping antigenic areas were defined by the anti-PA and anti...
Sequence analyses of the p24 gene of Borna disease virus in naturally infected horse, donkey and sheep. By reverse transcriptase/PCR amplification and subsequent sequence determination of the p24 gene, the relatedness of Borna disease virus (BDV) in various naturally infected animal species was determined. These results are indicative of a common ancestral virus pool and a remarkably low species barrier of BDV. Comparison of 11 sequences to that of tissue culture adapted virus revealed that the homology among all isolates was at least 96.2% at the nucleotide level, and 97% at the amino acid level. Viral sequences from sheep, donkey and horse were found to be not more distantly related to each ot...
A rapid method for the analysis of influenza virus genes: application to the reassortment of equine influenza virus genes. We describe a rapid method for genetic characterisation of influenza virus genes using reverse transcription and amplification by polymerase chain reaction (RT/PCR) of all virus segments simultaneously (multiplex RT/PCR) using primers based on the conserved terminal sequences. The product has been shown to be suitable for determination of partial nucleotide sequences which can be used to search nucleotide sequence databases and rapidly map the genetic origin of each segment. We illustrate the use of the method by analysing genetic reassortment in H7N7 equine influenza viruses.