Molecular Characterization, Sequence Analysis, and Taxonomic Position of Newly Isolated Fish Iridoviruses

Mao, Jinghe; Hedrick, Ronald P.; Chinchar, V. Gregory

doi:10.1006/viro.1996.8435

Cited by 283 publications

(329 citation statements)

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“…Confirmation of the relationship or divergence among systemic as well as between lymphocystis-inducing iridoviruses, in addition to visual evidences, e.g., dif- ferences in size and configuration, requires characterizing purified viral material by a variety of molecular approaches (Hedrick & McDowell 1995, Mao et al 1997. The iridovirus-like causative agents of systemic infections of fish, related by their properties with epizootic hematopoietic necrosis virus (EHNV) originally isolated from Perca fluviatilis in Australia (Langdon 1989), have been shown to have strong affinity with systemic amphibian viruses grouped into the genus Ranavirus .…”

Section: Discussionmentioning

confidence: 99%

Iridovirus infections in farm-reared tropical ornamental fish

Paperna¹,

Vilenkin²,

Matos³

2001

Dis. Aquat. Org.

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A systemic viral infection in both gourami Trichogaster spp. and swordtail Xiphophorus hellerii and an outbreak of lymphocystis in scalare Pterophyllum scalarae and gourami are reported to have occurred in fish reared in ornamental fish farms in Israel. The systemic infection developed in endothelial cells that became hypertrophic and their contents were modified. The presence of such cells in light-microscopically examined stained smears and sections provides an initial indication for this systemic viral infection. Infection in gourami caused hemorrhagic dropsy. Transmission electron microscopic (TEM) images of iridovirus-like particles recovered from gouramies showed them to be 138 to 201 nm from vertex to vertex (v-v); those from swordtails were 170 to 188 nm v-v. TEM images of lymphocystis virions from scalare were 312 to 342 nm v-v and from gourami 292 to 341 nm v-v. Lymphocystis cells from the gourami were joined by a solid hyaline plate, which was lacking in the infection in scalare where the intercellular spaces between the lymphocystis cells consisted of loose connective tissue.KEY WORDS: Trichogaster spp. · Xiphophorus spp. · Pterophyllum scalarae · Iridovirus-like particles · Systemic infection · Endothelium · Hypertrophy · Lymphocystis Resale or republication not permitted without written consent of the publisherDis Aquat Org 48: [17][18][19][20][21][22][23][24][25] 2001 findings together with a demonstration of viral particles by electron microscopy prove to be useful in the differential diagnosis of causes of mortality in the farmed fish. MATERIALS AND METHODSSmears and touch preparations were air-dried, fixed with absolute methanol and stained with Giemsa (in pH 7.4 buffer, for 1 h). For light microscopic histology, tissues were fixed in neutral-buffered formalin (exceptionally with aqueous Bouin's), and after dehydration in graded ethanols were embedded in glycol-methacrylate medium (Agar Scientific, Ltd, Stansted, UK). Sections, 2 to 3 µm, were cut with a glass knife on a Sorval JB4 microtome, and were stained with either Meyer's hemalum-eosin or, after a 20 min incubation in Bouin's solution and a water and 70% ethanol rinse, with 10% Giemsa in phosphate buffer, pH 7.4.For transmission electron microscopy (TEM), fragments from tissue with conspicuous cytopathological changes were fixed in 2.5% glutaraldehyde in cacodylate buffer (0.1 M, pH 7.4) for 24 h at 4°C, rinsed repeatedly in the same buffer, post-fixed in 1.0% osmium tetroxide in the same buffer for 1 h and, after rinsing in the buffer, dehydrated in graded ethyl alcohols and embedded in Agar 100 medium (Agar Scientific, Ltd). Thin sections, cut on a Reichert Ultracut Microtome with a diamond knife, were stained on grid with uranyl acetate and lead citrate, and examined with a Jeol 100CX TEM. Semi-thin sections cut with a glass knife were stained with Toluidine blue. RESULTS Viral hemorrhagic dropsy (VHD) in gouramiAffected fish, observed on several occasions in stocks of farmed gourami of the species Trichogaster trichopter...

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Section: Discussionmentioning

confidence: 99%

Iridovirus infections in farm-reared tropical ornamental fish

Paperna¹,

Vilenkin²,

Matos³

2001

Dis. Aquat. Org.

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“…As all of the samples were collected and processed in a similar manner, it may be assumed that most (or all) other DNA samples were of equal quality and that the negative PCR results accurately reflect the absence of virus in the samples, or its presence in such minute amounts that even an extremely sensitive test such as PCR could not detect it. Only 2/72 postmetamorphic green frogs thoroughly sampled in 2011 (blood, hepatic FNA, toe clip, and liver) were positive for ranavirus infection through PCR (Mao et al 1997). The liver was positive in both cases, whereas only one of the frogs had a positive toe-clip (Table 1).…”

Section: Discussionmentioning

confidence: 99%

“…Total DNA was extracted from all samples using a spin-column DNA purification procedure (Qiagen DNeasy 96, Qiagen, Valencia, California, USA). The blood, toe clips, hepatic FNA, and liver and kidney tissue samples were tested for the ranavirus major capsid protein gene with single-round PCR amplification (Mao et al, 1997), using primers covering the same region of the MCP gene as the MCP1 assay recommended by the Aquatic Animal Health Code (World Organisation for Animal Health, 2012).…”

Section: Methodsmentioning

confidence: 99%

LOW DETECTION OF RANAVIRUS DNA IN WILD POSTMETAMORPHIC GREEN FROGS, RANA (LITHOBATES) CLAMITANS, DESPITE PREVIOUS OR CONCURRENT TADPOLE MORTALITY

Forzán

Wood

2013

Journal of Wildlife Diseases

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ABSTRACT:Ranavirus (Iridoviridae) infection is a significant cause of mortality in amphibians. Detection of infected individuals, particularly carriers, is necessary to prevent and control outbreaks. Recently, the use of toe clips to detect ranavirus infection through PCR was proposed as an alternative to the more frequently used lethal liver sampling in green frogs (Rana [Lithobates] clamitans). We attempted reevaluate the use of toe clips, evaluate the potential use of blood onto filter paper and hepatic fine needle aspirates (FNAs) as further alternatives, and explore the adequacy of using green frogs as a target-sampling species when searching for ranavirus infection in the wild. Samples were obtained from 190 postmetamorphic ($1-yr-old) green frogs from five ponds on Prince Edward Island (PEI), Canada. Three of the ponds had contemporary or recent tadpole mortalities due to Frog Virus 3 (FV3) ranavirus. PCR testing for ranavirus DNA was performed on 190 toe clips, 188 blood samples, 72 hepatic FNAs, and 72 liver tissue samples. Only two frogs were ranavirus-positive: liver and toe clip were positive in one, liver only was positive in the other; all blood and FNAs, including those from the two positive frogs, were negative. Results did not yield a definitive answer on the efficacy of testing each type of sample, but resemble what is found in salamanders infected with Ambystoma tigrinum (rana)virus. Findings indicate a low prevalence of FV3 in postmetamorphic green frogs on PEI (#2.78%) and suggest that green frogs are poor reservoirs (carriers) for the virus.

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“…The hepatic tissue samples (2-3 mm 3 ) were handled according to Duffus et al (2008) for DNA extraction. The DNA extract was screened for the presence of the MCP of FV3 following Mao et al (1997) using primer set MCP 4/5. The PCR products were then run on a 1.5% ethidium bromide-stained agarose gel to determine the presence of the MCP gene by the appearance of an approximately 500 base pair (bp) band.…”

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confidence: 99%