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At least 19 recordsLinked to original sources
PubMed2001

Comparison of the eIF-2alpha homologous proteins of seven ranaviruses (Iridoviridae).

The alpha-subunit of the eukaryotic initiation factor 2 (eIF-2alpha) is a key component of the translation machinery of the cell. In response to cellular stress such as viral infections, eIF-2alpha is phosphorylated by double-stranded RNA-dependent protein kinase (PKR) leading to the inhibition of cellular protein synthesis. The importance of eIF-2alpha as a regulatory mechanism for protein synthesis is illustrated by the wide variety of strategies employed by viruses to down-regulate PKR. Thus, Vaccinia virus encodes K3L protein, which resembles eIF-2alpha and acts as a pseudo-substrate inhibitor of PKR. Nucleotide sequencing of the genome of epizootic haematopoietic necrosis virus (EHNV), a member of the genus ranavirus of Iridoviridae, has revealed an eIF-2alpha equivalent gene. We have cloned and sequenced eIF-2alpha genes of several iridoviruses of fishes and frogs. The eIF-2alpha open reading frames and deduced proteins of the iridoviruses investigated exhibit a high degree of homology of both nucleotide and amino acid sequences. At the N-terminus, the iridoviral eIF-2alpha shows significant homology to the N-termini of cellular initiation factor 2-alpha of various species, to full-length poxviral eIF-2alpha proteins, and to the S1 domain of ribosomal proteins. Comparison of amino acid sequences of corresponding iridoviral proteins with eIF-2alpha homologous proteins of poxviruses and eukaryotes has revealed a high conservation of motifs. A phylogenetic analysis of eukaryotic eIF-2alpha and poxvirus and iridovirus eIF-2alpha sequences has demonstrated the relationship of these iridoviruses. In order to investigate the role of the eIF-2alpha equivalent, respective genes have been expressed in prokaryotic and eukaryotic (insect, fish and chicken cell) systems. The iridoviral eIF-2alpha protein has a molecular weight of 31 kDa and is cytoplasmic. The cellular and viral protein synthesis of iridoviruses is probably regulated by a mechanism similar to that of Vaccinia virus. Frog-virus 3, the type species of the genus ranavirus of Iridoviridae, has a unique translational efficiency and, moreover, down-regulates the cellular protein synthesis of infected cells.

Amino Acid Sequence↗
PubMed2004

Comparative genomic analyses of frog virus 3, type species of the genus Ranavirus (family Iridoviridae).

Frog virus 3 (FV3) is the type species member of the genus Ranavirus (family Iridoviridae). To better understand the molecular mechanisms involved in the replication of FV3, including transcription of its highly methylated DNA genome, we have determined the complete nucleotide sequence of the FV3 genome. The FV3 genome is 105903 bp long excluding the terminal redundancy. The G + C content of FV3 genome is 55% and it encodes 98 nonoverlapping potential open reading frames (ORFs) containing 50-1293 amino acids. Eighty-four ORFs have significant homology to known proteins of other iridoviruses, whereas twelve of these unique FV3 proteins do not share homology to any known protein. A microsatellite containing a stretch of 34 tandemly repeated CA dinucleotide in a noncoding region was detected. To date, no such sequence has been reported in any animal virus.

Animals↗
PubMed2004

Investigations on the ORF 167L of lymphocystis disease virus (Iridoviridae).

The predicted open reading frame 167L (ORF 167L) of lymphocystis disease virus (LCDV, Iridoviridae ) isolated from plaice, dab and flounder was investigated. The ORF 167L corresponding genes of the three LCDV isolates were amplified, cloned and sequenced. A comparison of the LCDV strains showed that the nucleotide sequence of ORF 167L and its deduced amino acid sequence were highly conserved in the genus lymphocystivirus (a homology of 80% in dab and flounder/plaice, 97% in plaice and flounder). The N-terminus protein predicted from the ORF 167L suggests similarities to the tumor necrosis factor receptor (TNFR)-family, and to TNFR-like proteins, which play an important role in various poxvirus species. Further, homology to the CUB-domain was shown at the C-terminus of the LCDV protein. Phylogenetic analyses of partial LCDV protein sequences identified two clusters: one cluster containing the flounder and plaice LCDV isolate (LCDV-1), and another cluster, containing the dab LCDV isolate (LCDV-2). The ORF 167L of plaice LCDV was expressed in Escherichia coli, and in fish cells. The expressed ORF resulted in a 30-kDa cytoplasmic protein lacking a signal peptide. An established monoclonal antibody (mAb 18) was used to detect LCDV proteins in skin explants of flounders and cryosections of dab skin. Specific fluorescence was found in the cytoplasm of intact epitheloid cells of the lymphocystis capsule and in the epidermis skin covering the lymphocystic nodules. LCDV-specific labelling of mAb 18 was also shown in spleen and liver tissue of LCDV-positive flounders. The ORF 167L protein seemed not to have the extracellular receptor function predicted from the usual cellular TNFR. The myxomavirus M-T2 protein, a poxviral TNFR homologue, was also shown not to have TNFR-like functions but to be involved in the apoptosis signal cascade.

Amino Acid Sequence↗
PubMed1995

Identification and properties of the largest subunit of the DNA-dependent RNA polymerase of fish lymphocystis disease virus: dramatic difference in the domain organization in the family Iridoviridae.

Cytoplasmic DNA viruses encode a DNA-dependent RNA polymerase (DdRP) that is essential for transcription of viral genes. The amino acid sequences of the known largest subunits of DdRPs from different species contain highly conserved regions. Oligonucleotide primers, deduced from two conserved domains (RQP[T/S]LH and NADFDGDE) were used for detecting the corresponding gene of fish lymphocystis disease virus (FLCDV), a member of the family Iridoviridae, which replicates in the cytoplasm of infected cells of flatfish. The gene coding for the largest subunit of the DdRP was identified using a PCR-derived probe. The screening of the complete EcoRI gene library of the viral genome led to the identification of the gene locus of the largest subunit of the DdRP within the EcoRI DNA fragment B (12.4 kbp, 0.034 to 0.165 map units). The nucleotide sequence of a part (8334 bp) of the EcoRI DNA fragment B was determined and a large ORF on the lower strand (ATG = 5787; TAA = 2190) was detected which encodes a protein of 1199 amino acids. Comparison of the amino acid sequences of the largest subunits of the DdRP (RPO1) of FLCDV and Chilo iridescent virus (CIV) revealed a dramatic difference in their domain organization. Unlike the 1051 aa RPO1 of CIV, which lacks the C-terminal domain conserved in eukaryotic, eubacterial and other viral RNA polymerases, the 1199 aa RPO1 of FLCDV is fully collinear with its cellular and viral homologues. Despite this difference, comparative analysis of the amino acid sequences of viral and cellular RNA polymerases suggests a common origin for the largest RNA polymerase subunits of FLCDV and CIV.

Amino Acid Sequence↗
PubMed2003

Structural analyses of Phycodnaviridae and Iridoviridae.

The Phycodnaviridae, Iridoviridae and related viruses, with diameters of 1500-2000 A, are formed from large trigonal arrays of hexagonally close-packed capsomers forming the faces of icosahedra [Yan et al. (2000), Nature Struct. Biol. 7, 101-103; Nandhagopal et al. (2002), Proc. Natl Acad. Sci. USA, 99, 14758-14763]. Caspar and Klug predicted that such structures could be assembled from hexameric capsomers [Caspar & Klug (1962), Cold Spring Harbor. Symp. Quant. Biol. 27, 1-24], as was subsequently found in numerous icosahedral viruses. During the course of evolution, some viruses, including the virus families mentioned above, replaced hexameric capsomers with pseudo-hexameric trimers by gene duplication. In large dsDNA icosahedral viruses, the capsomers are organized into 'pentasymmetrons' and 'trisymmetrons'. The interactions between the trimeric capsomers can be divided into three groups, one between similarly oriented trimers and two between oppositely oriented trimers (trimers related by an approximately sixfold rotation). The interactions within a trisymmetron belong to the first class, whereas those between trisymmetrons and within the pentasymmetron are of the other two types. Knowledge of these distances permits a more accurate fitting of the atomic structure of the capsomer into the cryo-electron microscopy (cryoEM) reconstruction of the whole virus. The adoption of pseudo-hexagonal capsomers places these viruses into a subset of the Caspar and Klug surface lattices.

Capsid Proteins↗
PubMed2002

Ranaviruses (family Iridoviridae): emerging cold-blooded killers.

Although possessing novel replicative and structural features, the family Iridoviridae has not been as extensively studied as other families of large, DNA-containing viruses (e.g., poxviridae and herpesviridae). This oversight most likely reflects the inability of iridoviruses to infect mammals and birds, and their heretofore low pathogenicity among cold-blooded animals and invertebrates. In fact, the original frog virus isolates (e.g., frog viruses 1-3) would likely have been considered orphan viruses since they were isolated from apparently healthy frogs. However, recent disease outbreaks among commercially and recreationally important fish, cultured and wild frogs, and endangered salamanders has challenged this benign view and have implicated several members of the genus Ranavirus as pathogens. This review explores three facets of ranavirus biology. In the first the salient features of ranavirus replication are summarized using frog virus 3 as a model. Secondly, criteria for characterizing new ranavirus isolates, based on biochemical (viral protein profiles, DNA restriction fragment length polymorphisms, and nucleotide sequence analysis), ecological (host range, tissue tropism), and clinical considerations, are detailed. Lastly, the principal agents of ranavirus-mediated disease and immune responses to these viruses are discussed. In light of the above, it is clear that ranaviruses are no longer orphan viruses, and that they have a significant impact on diverse populations of ectothermic animals.

Amphibians↗
PubMed2003

Genomic sequence of a ranavirus (family Iridoviridae) associated with salamander mortalities in North America.

Disease is among the suspected causes of amphibian population declines, and an iridovirus and a chytrid fungus are the primary pathogens associated with amphibian mortalities. Ambystoma tigrinum virus (ATV) and a closely related strain, Regina ranavirus (RRV), are implicated in salamander die-offs in Arizona and Canada, respectively. We report the complete sequence of the ATV genome and partial sequence of the RRV genome. Sequence analysis of the ATV/RRV genomes showed marked similarity to other ranaviruses, including tiger frog virus (TFV) and frog virus 3 (FV3), the type virus of the genus Ranavirus (family Iridoviridae), as well as more distant relationships to lymphocystis disease virus, Chilo iridescent virus, and infectious spleen and kidney necrosis virus. Putative open reading frames (ORFs) in the ATV sequence identified 24 genes that appear to control virus replication and block antiviral responses. In addition, >50 other putative genes, homologous to ORFs in other iridoviral genomes but of unknown function, were also identified. Sequence comparison performed by dot plot analysis between ATV and itself revealed a conserved 14-bp palindromic repeat within most intragenic regions. Dot plot analysis of ATV vs RRV sequences identified several polymorphisms between the two isolates. Finally, a comparison of ATV and TFV genomic sequences identified genomic rearrangements consistent with the high recombination frequency of iridoviruses. Given the adverse effects that ranavirus infections have on amphibian and fish populations, ATV/RRV sequence information will allow the design of better diagnostic probes for identifying ranavirus infections and extend our understanding of molecular events in ranavirus-infected cells.

Animals↗
PubMed2001

Occurrence of an invertebrate iridescent-like virus (Iridoviridae) in reptiles.

Viral isolates were obtained in 1998, 1999 and 2000 from the lung, liver and intestine of two bearded dragons (Pogona vitticeps) and a chameleon (Chamaeleo quadricornis) and from the skin of a frill-necked lizard (Chamydosaurus kingii) by using viper heart cells (VH2) at 28 degrees C. Electron microscopic examination of infected VH2 cells revealed the assembly of icosahedral iridovirus-like particles measuring 139 nm (side to side) and 151 nm (apex to apex). Negatively stained virus particles had dimensions of 149 nm (side to side) and 170 nm (apex to apex). Polymerase chain reaction (PCR) amplification of purified viral DNA with primers corresponding to the partial gene encoding the major capsid protein (MCP) of Frog viris-3 (FV-3), the type species of the genus Ranavirus, was unsuccessful. In contrast, primers corresponding to the partial MCP gene of Chilo iridescent virus (CIV; genus Iridovirus) amplified 500-bp products with 97% identity to the nucleotide sequence of CIV and 100% identity to the nucleotide sequence of Gryllus bimaculatus iridescent virus (GbIV), an invertebrate iridescent virus. Virus protein profiles analysed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and restriction fragment length profiles of purified viral DNA treated with the endonucleases EcoRI, HindIII and HpaII were identical to those of GbIV.

Animals↗
PubMed2002

The epizootic haematopoietic necrosis virus (Iridoviridae) induces apoptosis in vitro.

The epizootic haematopoietic necrosis virus (EHNV) is an iridovirus causing severe disease in different fish species. We investigated the induction of apoptosis during EHNV infection of the epithelioma carp papulosum (EPC) cell line. Apoptosis reveals several characteristic morphological changes, such as chromatin condensation, nuclear fragmentation, cytoplasm membrane disorientation, or mitochondrial changes. During EHNV infection of EPC cells the occurrence of apoptosis was analysed using a fluorescein-isothiocyanate (FITC) conjugate of annexin-V to detect phosphatidylserines that have changed cytoplasm membrane localization. Annexin-V labelling was obvious 12 h after infection. At 54 h after EHNV infection 39% of the investigated EPC cells exhibited fluorescence. Furthermore, EHNV-infected cells were stained with 4'-6'-diamidino-2-phenylindole (DAPI) to detect pycnotic nuclei. Appearance of DAPI-positive nuclei was found beginning at 18 h after infection. At 54 h after EHNV infection approximately 56% of the EPC cells showed fragmented nuclei. Assays to inhibit a protein kinase-dependent (e.g. double-stranded RNA-dependent protein kinase) apoptosis pathway with 2-aminopurine revealed a reduction of EHNV titres, e.g. titres were reduced 1000-fold in the presence of 100 and 200 mM 2-aminopurine. Apoptosis takes place during iridovirus infection in vitro and it seems to involve the activation of protein kinases.

Animals↗
PubMed1997

Development of DNA diagnostic methods for the detection of new fish iridoviral diseases.

A new disease of epidemic proportions caused by fish viruses within the Iridoviridae family inflicts serious damage on red sea breams (Pagrus major) and striped jack (Caranx delicatissimus) populations grown in aquacultures in Japan. A partial segment of the fish iridoviral DNA was directly amplified using the polymerase chain reaction (PCR) with synthetic primers designed from well conserved nucleotide sequences between the frog virus 3 (Ranavirus) and the silkworm iridescent virus type 6. The deduced amino acid sequence from the nucleotide sequence of the PCR fragment demonstrates a high correlation with a partial sequence from the frog virus 3. Using the PCR method with specific primers, we could detect three of four different known types of fish iridoviruses in diseased fishes. To construct more reliable detection methods specific for this viral family, DNA fragments which can specifically hybridize with all of the four known iridoviridae viral DNAs were screened from the genomic library of one iridoviridae strain. The hybridization assay, using a specific fragment which contains regions which are highly homologous with a characterized partial sequence from the frog virus 3, proved to be a reliable diagnostic tool for fish iridoviral diseases.

Animals↗
PubMed2006

Intracytoplasmic inclusions in circulating leukocytes from an eastern box turtle (Terrapene carolina carolina) with iridoviral infection.

A free-ranging adult female eastern box turtle (Terrapene carolina carolina) was presented to the University of Tennessee in October 2003 because of suspected trauma and blindness. Physical examination revealed lethargy, clear ocular and nasal discharges, and white oral and laryngeal plaques. Intracytoplasmic inclusions within heterophils and large mononuclear leukocytes were observed on routine blood smear examination. Postmortem findings included necrosis of epithelial and parenchymal cells with intracytoplasmic inclusions. Ultrastructurally, the leukocyte inclusions consisted of variably electron-dense granular material and viral particles consistent with the Iridoviridae family of viruses. The virus shared 100% sequence identity to a 420-base pair sequence of frog virus 3 (family Iridoviridae, genus Ranavirus) as determined by polymerase chain reaction and gene sequencing targeting a portion of the Ranavirus major capsid protein gene.

Animals↗
PubMed2002

Viral DNA sequences of genes encoding the ATPase and the major capsid protein of tropical iridovirus isolates which are pathogenic to fishes in Japan, South China Sea and Southeast Asian countries.

Tropical iridovirus infection causes severe epizootic resulting in mass mortalities and large economic losses in freshwater ornamental fishes cultured in Southeast Asian countries, in wild fish seedlings captured in South China Sea, and in marine fishes farmed in Japan, Singapore, and Thailand. All of tropical iridovirus-infected fishes histopathologically showed the systemic formation of inclusion body-bearing cells and necrosis of virus-infected splenocytes and hematopoietic cells. We designed primer sets for the ATPase gene and the major capsid protein (MCP) gene and sequenced the PCR products derived from 5 iridovirus isolates from sea bass in South China Sea, red sea bream in Japan, brown-spotted grouper with a grouper sleepy disease in Thailand, dwarf gourami from Malaysia and African lampeye from Sumatra Island, Indonesia. The ATPase gene and the MCP gene of these 5 viral isolates were highly homologous (> 95.8%, > 94.9% identity, respectively) and the deduced amino acid sequences of the ATPase and the MCP were also highly identical (> 98.1%, > 97.2% identity, respectively). Based on the high homology, these 5 isolates of tropical iridovirus from various fishes in geographically different regions were determined to have a single origin and to be native to Southeast Asian regions. However, these sequences were far different from those of members of the genera Ranavirus, Lymphocystivirus and Iridovirus in the Family Iridoviridae. We propose a new genus "Tropivirus" for tropical iridovirus in the Family Iridoviridae.

Adenosine Triphosphatases↗
PubMed2001

Transcription and temporal cascade in Chilo iridescent virus infected cells.

Chilo iridescent virus (CIV) is the type species for genus Iridovirus, and belongs to the family Iridoviridae. Members of this family are large, isometric, cytoplasmic DNA viruses. Our laboratory has established that CIV replicates productively in the cotton boll weevil, Anthonomus grandis. Given the economic importance of this host and the dearth of knowledge on this virus, we have initiated host-virus interaction and molecular studies on CIV. This report focuses on regulation of transcription in CIV infections. We carried out northern analyses on total cellular RNA from infections of IPRI-CF-124T cells, using a complete genomic library of CIV and several putative gene-specific probes. Our data show a temporal cascade based on analysis of 137 detectable transcripts comprising 38 immediate-early (IE), 34 delayed-early (DE), and 65 late (L) transcripts. Analysis with gene-specific probes supported the cascade pattern. Both helicase and RNA polymerase were immediate-early; major capsid protein was late. The CIV gene expression cascade appears to operate primarily at the transcriptional level. Temporal classes observed are consistent with earlier studies at the polypeptide level and with transcriptional patterns in frog virus 3, genus Ranavirus in the Iridoviridae. Our results provide an important basis for understanding mechanisms driving the CIV temporal cascade.

Aphidicolin↗
PubMed2003

Molecular characterization of the major virion protein gene from the Trichoplusia ni ascovirus.

Ascoviruses (AVs) belong to a family of double-stranded DNA viruses that infect Lepidoptera insects and cause the unique pathology of forming virion-containing vesicles in the hemolymph of infected hosts. Virions of AVs are large and contain more than 12 polypeptides. A gene, TnAV-CP, encoding the major structural protein of the Trichoplusia ni ascovirus 2a (TnAV-2a) was cloned by immunoscreening an expression library with antibodies against total TnAV virion proteins. TnAV-CP is an intronless gene with an open reading frame encoding a protein of 455 amino acids. Southern blot showed that it is a single copy gene. A 3.8 kb BamHI fragment containing the complete TnAV-CP gene was cloned and sequenced. Northern analysis detected the transcription of the 1.4 kb TnAV-CP mRNA from 24 h after infection. The predicted TnAV-CP protein was expressed in bacterial expression system and purified to homogeneity. The recombinant protein was used to affinity-purify specific antibodies from the antiserum. The purified antibodies reacted strongly with a single protein of approximately 52 kDa from the total TnAV virion proteins in a Western blot. This protein corresponds to the most abundant structural protein present in the virions of several AVs. Sequence comparison showed that TnAV-CP is most homologous to the putative capsid proteins from AVs infecting noctuid insects, less homologous to that of Diadromus pulchellus ascovirus 4a (DpAV-4a), further supporting the distinction of two subgroups within the family Ascoviridae. Phylogenetic analysis using the putative capsid protein suggested that AVs were closely related to members of Iridoviridae, which corroborated the result based on DNA polymerase delta sequences. The apparent differences between Ascoviridae and Iridoviridae in host range, virion morphology, and genome configuration, and the similarities in genes and methylation of genomic DNA were discussed.

Amino Acid Sequence↗
PubMed2002

Comparative analysis of the genome and host range characteristics of two insect iridoviruses: Chilo iridescent virus and a cricket iridovirus isolate.

The iridovirus isolate termed cricket iridovirus (CrIV) was isolated in 1996 from Gryllus campestris L. and Acheta domesticus L. (both Orthoptera, Gryllidae). CrIV DNA shows distinct DNA restriction patterns different from those known for Insect iridescent virus type 6 (IIV-6). This observation led to the assumption that CrIV might be a new species within the family Iridoviridae. CrIV can be transmitted perorally to orthopteran species, resulting in specific, fatal diseases. These species include Gryllus bimaculatus L. and the African migratory locust Locusta migratoria migratorioides (Orthoptera, Acrididae). Analysis of genomic and host range properties of this isolate was carried out in comparison to those known for IIV-6. Host range studies of CrIV and IIV-6 revealed no differences in the peroral susceptibility in all insect species and developmental stages tested to date. Different gene loci of the IIV-6 genome were analyzed, including the major capsid protein (274L), thymidylate synthase (225R), an exonuclease (012L), DNA polymerase (037L), ATPase (075L), DNA ligase (205R) and the open reading frame 339L, which is homologous to the immediate-early protein ICP-46 of frog virus 3. The average identity of the selected viral genes and their gene products was found to be 95.98 and 95.18% at the nucleotide and amino acid level, respectively. These data led to the conclusion that CrIV and IIV-6 are not different species within the Iridoviridae family and that CrIV must be considered to be a variant and/or a novel strain of IIV-6.

Amino Acid Sequence↗
PubMed2002

Sequence analysis of the complete genome of an iridovirus isolated from the tiger frog.

We have isolated a tiger frog virus (TFV) from diseased tiger frogs, Rana tigrina rugulosa. The genome was a linear double-stranded DNA of 105,057 basepairs in length with a base composition of 55.01% G+C. About 105 open reading frames were identified with coding capacities for polypeptides ranging from 40 to 1294 amino acids. Computer-assisted analyses of the deduced amino acid sequences revealed that 39 of 105 putative gene products showed significant homology to functionally characterized proteins of other species in the GenBank/EMBL/DDBJ databases. These proteins included enzymes and structural proteins involved in virus replication, transcription, modification, and virus--host interaction. The deduced amino acid sequences of TFV gene products showed more than 90% identity to FV3, but a low degree of similarity among TFV, ISKNV, and LCDV-1. The results from this study indicated that TFV may belong to the genus Ranavirus of the family Iridoviridae.

Amino Acid Sequence↗
PubMed1997

Molecular anatomy of lymphocystis disease virus.

Lymphocystis disease (LD) has been reported to occur in over one hundred different species of fish worldwide. The disease is caused by lymphocystis disease virus (LCDV), a member of the iridovirus family. Numerous fish species that play an important role in fishery and fish farming are highly susceptible to LCDV infection. The infected animals develop disseminated clusters of aberrant hypertrophied cells within their connective tissue, the so-called lymphocystis cells. In the cytoplasm of these cells a massive accumulation of virions can be observed. As a first step towards understanding the mechanisms of viral infection and pathogenesis the complete genomic nucleotide sequence of lymphocystis disease virus type 1 (LCDV-1; flounder isolate) was determined. LCDV-1 is the type species of the genus Lymphocystivirus within the family Iridoviridae. The virions contain a single linear double-stranded DNA molecule that is circularly permuted, terminally redundant and heavily methylated. Since there is no convenient cell system for virus replication we determined the complete nucleotide sequence of the viral genome (102,653 base pairs). Computer assisted analyses of 195 potential open reading frames resulted in the identification of a number of putative gene products with significant homology to functionally characterized proteins of other species.

Animals↗