Epizootic Haematopoietic Necrosis Virus Research Articles

Purification of epizootic haematopoietic necrosis virus and its detection using ELISA

Epizootic haematopoietic necrosis virus (EHNV) was grown in Bluegill fry (BF-2) cells and purified using differential and gradient centrifugation. The lower band (B2) from 15–60% sucrose gradients contained infective EHNV but few contaminating cell components when assessed by electron microscopy, SDS-PAGE, and Western blotting using anti-BF-2 serum and anti-B2 serum. Both rabbit and sheep anti-B2 sera precipitated B2 in agarose gel immunodiffusion and detected EHNV in cell culture supernatant when used in an indirect antigen-capture ELISA. Rabbit anti-B2 serum was used as capture antibody while sheep anti-B2 serum was used to detect viral antigen. Pre-adsorption of diluted sheep anti-B2 serum using BF-2 cell lysate greatly improved the specificity and sensitivity of the technique.

Epizootic haematopoietic necrosis virus: purification and classification

Abstract. Epizootic haematopoietic necrosis virus (EHNV) has been isolated from redfin perch, Perca fluviatilis L., and cultured rainbow trout, Oncorhynchus mykiss (Walbaum), in Victoria and New South Wales, Australia. On the basis of virion size, icosahedral morphology and ultrastructural characteristics of virus‐infected cells, EHNV was designated as an iridovirus. The following observations outlined in this paper confirm the designation: (1) The number and complexity of proteins in purified EHNV and the ordered appearance of virus‐specific proteins in infected cells are similar to that described for the iridovirus, frog virus 3 (FV3). (2) EHNV isolated from the cytoplasm of infected cells contains a lipid membrane associated with 7–9 nm capsomeric subunits containing a 52‐kD protein. (3) The morphogenesis of EHNV within cytoplasmic inclusion bodies resembles that of FV3 and a piscine iridovirus isolated from goldfish, Carassius auratus (L.) (4) EHNV also resembles FV3 in its association with the cytoskeleton of infected cells. (5) EHNV DNA, like that of other iridoviruses, is resistant to digestion with HpaII, a restriction endonuclease which does not cleave DNA at CCGG sequences where the internal cytosine is methylated.

Experimental transmission and pathogenicity of epizootic haematopoietic necrosis virus (EHNV) in redfin perch, Perca fluviatilis L., and 11 other teleosts

Abstract. The epizootiology of epizootic haematopoietic necrosis virus (EHNV) was examined by experimental transmission and pathogenicity trials in the first known host species, Perca fluviatilis L., and 11 other introduced and native teleosts of Australia. Perca fluviatilis and four other species were highly susceptible to disease following bath exposure to EHNV. Three species developed disease only when inoculated by intraperitoneal injection, and four species wore resistant to infection under the experimental conditions employed. Renal haematopoietic necrosis was the most consistent lesion produced, with hepatic, splenic and pancreatic necrosis also occurring in certain species. The resistance of the virus to desiccation, disinfectants, high and low pH and temperature, and to prolonged storage at selected temperatures was assessed, demonstrating that the virus can survive for long periods in the aquatic environment and on fomites. Carrier individuals of P. fluviatilis, Salmo gairdneri Richardson, Maccullochella peeli (Mitchell), and possibly other teleosts, represent a further reservoir of infection. However, the virus could not be isolated from common insect and crustacean prey species, and no parasitic vectors were identified. It is proposed that EHNV has contributed to serious population declines of native species in recent decades, particularly of Macquaria australasica (Cuvier), Galaxias olidus Günther and Bidyanus bidyanus (Mitchell). Salmonids are also known to be naturally and experimentally susceptible to the virus.

Spinal curvatures and an encephalotropic myxosporean, Triangula percae sp. nov. (Myxozoa: Ortholineidae), enzootic in redfin perch, Perca fluviatilis L., in Australia

Abstract. A new myxosporean, Triangula percae sp. nov., (Myxozoa: Ortholineidae) from the brain of redfin perch, Perca fluviatilis L., is described. Spinal curvatures are prevalent in many perch populations in Victoria, Australia, Triangula percae infection was detected in all perch displaying spinal curvatures and in 67% of normal perch from Lake Nillahcootie in north east Victoria. Perch populations exhibiting spinal curvatures in several other waterways were also infected with Triangula percae but the infection was absent in one population which was apparently normal. Triangula percae infection is, thus, considered to be the cause of a proportion of the spinal curvatures. Certain fish surviving outbreaks of epizootic haematopoietic necrosis virus (EHNV) also developed spinal curvatures, which may have been due to viral infection of the central nervous system.