Study of the viral interference between infectious pancreatic necrosis virus (IPNV) and infectious haematopoietic necrosis virus (IHNV) in rainbow trout (Oncorhynchus mykiss)

The resistance of rainbow trout (Oncorhynchus mykiss) to an infectious haematopoietic necrosis virus (IHNV) challenge following a preceding non-lethal infection with infectious pancreatic necrosis virus (IPNV) was investigated through experimental dual infections. Trout initially infected with IPNV were inoculated 14 days later with IHNV. Single infections of trout with 1 of the 2 viruses or with cell culture supernatant were also carried out and constituted control groups. No mortality was noted in fish after a single infection with IPNV. This virus had no influence on the head kidney leucocyte phagocytic activity and plasma haemolytic complement activity. IHNV induced a high mortality (72%) and reduced the macrophage phagocytic activity and complement haemolytic activity. It also induced a late production of anti-IHNV antibodies which occurred after clearance of the virus in the fish. In trout co-infected with both viruses, a mortality rate of 2% occurred and the immune parameters were similar to those observed in the fish infected with IPNV only, demonstrating that in co-infected trout IPNV inhibits the effects of IHNV. The studied parameters did not allow us to define the mechanism of interference occurring between these 2 viruses, but some hypothesis are put forward to explain the interference between the 2 viruses.     

First isolation of an aquatic birnavirus from farmed and wild fish species in Australia

During routine sampling and testing, as part of a systematic surveillance program (the Tasmanian Salmonid Health Surveillance Program), an aquatic birnavirus was isolated from 'pin-head' (fish exhibiting deficient acclimatisation on transfer to saltwater) Atlantic salmon Salmo salar, approximately 18 mo old, farmed in net-pens located in Macquarie Harbour on the west coast of Tasmania, Australia. The isolate grows readily in a range of fish cell lines including CHSE-214, RTG-2 and BF-2 and is neutralised by a pan-specific rabbit antiserum raised against infectious pancreatic necrosis virus (IPNV) Ab strain and by a commercial pan-specific IPNV-neutralising monoclonal antibody. Presence of the virus was not associated with gross clinical signs. Histopathological examination revealed a range of lesions particularly in pancreatic tissue. The virus was localised in pancreas sections by immunoperoxidase staining using the polyclonal antiserum and by electron microscopy. Examination by electron microscopy demonstrated that the virus isolated in cell culture (1) belongs to the family Birnaviridae, genus Aquabirnaviridae; (2) was ultrastructurally and antigenically similar to virus identified in the index fish; (3) is related to IPNV. Western blot analysis using the polyclonal rabbit antiserum confirmed the cross-reactions between various aquatic birnavirus isolates. In addition, PCR analysis of isolated viral nucleic acid from the index case indicated that the virus is more closely related to IPNV fr21 and N1 isolates than to other birnavirus isolates available for comparison. Sampling of other fish species within Macquarie Harbour has demonstrated that the virus is present in several other species of fish including farmed rainbow trout Oncorhynchus mykiss, wild flounder Rhombosolea tapirina, cod Pseudophycis sp., spiked dogfish Squalus megalops and ling Genypterus blacodes.     

Co-occurrence of viral and bacterial pathogens in disease outbreaks affecting newly cultured sparid fish.

Several microbial disease outbreaks in farm stocks of newly cultured sparid fish species, such as common seabream, redbanded seabream, and white seabream, were recorded from 2004 to 2006. This study describes the isolation and characterization of the potential causative agents, either bacteria or viruses, of these outbreaks. The isolated bacterial strains were characterized according to traditional taxonomical analyses and sequencing of a 16S rDNA fragment. Most bacteria were identified as Vibrio spp. and Photobacterium damselae subsp. damselae. The development of cytopathic effects (CPE) on different fish cell lines, the application of specific nested-PCR tests for infectious pancreatic necrosis virus (IPNV), viral nervous necrosis virus (VNNV) and viral hemorrhagic septicemia virus (VHSV), and subsequent sequence analyses were used for virus detection and identification. VNNV, related to the striped jack neural necrosis virus (SJNNV) genotype, and VHSV, related to the genotype Ia, were the only viruses detected. VNNV was isolated from the three fish species under study in five different outbreaks, whereas VHSV was isolated from common seabream and white seabream during two of these outbreaks. IPNV was not detected in any case.     

Flow cytometry detection of infectious pancreatic necrosis virus (IPNV) within subpopulations of Atlantic salmon (Salmo salar L.) leucocytes after vaccination and during the time course of experimental infection

In the present study, intracellular infectious pancreatic necrosis virus (IPNV) in salmon leucocytes was detected by flow cytometry after experimental cohabitant challenge. IPNV vaccinated, non-vaccinated and intraperitoneally (i.p.) infected salmon (virus shedders) were analysed at different times throughout the period when mortality occurred. Fish that had survived 61 days post challenge (carriers) were also analysed. In particular, we analysed the presence of IPNV in B-cells (C7G7+cells) and in neutrophils (E3D9+ cells) in head kidney leucocytes (HKL) and in peripheral blood leucocytes (PBL).IPNV was present in HKL and PBL from all challenged fish groups at all samplings, including carriers. IPNV was also found intracellular in other leucocytes than B-cells and neutrophils. During the time course of infection there were changes in proportion of B-cells and neutrophils and in proportions of IPNV+ cells. In vaccinated fish, a delay in the changes observed in the proportion of IPNV+ cells and in the proportions of the two subpopulations was identified. The vaccinated fish were protected against disease as no fish died compared to 30.8% of non-vaccinated cohabitant fish. All i.p. infected fish, except one, survived the challenge. This is consistent with previous studies and confirmed that the routes of infection can influence mortality. The analyses in this study could not identify any factors enlightening this absence of mortality in i.p. infected fish, but both flow cytometry and qRT-PCR showed that i.p. infected fish were carriers of IPNV. The present study also found that IPNV was present in both B-cells and neutrophils as well as in other leucocytes in all carriers after cohabitant challenge. These fish had survived 9 weeks post challenge and 4 weeks after mortality has ceased. The fish harbouring virus within their leucocytes might become life long carriers and represent a risk for disease outbreaks, being virus shedders. Such fish are protected from later infections if the virus exposure has resulted in protective immunity. Flow cytometry was found to be very suitable for detection of intracellular virus after in vivo challenge and the sensitivity was demonstrated by the detection of virus in carriers.     

Infectious pancreatic necrosis virus infection in Atlantic salmon Salmo salar post-smolts affects the outcome of secondary infections with infectious salmon anaemia virus or Vibrio salmonicida.

Infectious pancreatic necrosis (IPN) virus (IPNV) infection in Atlantic salmon Salmo salar L. post-smolts and its influence on the outcome of secondary infections with infectious salmon anaemia (ISA) virus (ISAV) or Vibrio salmonicida were studied. The infections with ISAV or V salmonicida were performed both in a period of acute IPN and in the following IPNV carrier stage, 3 and 6 to 8 wk after experimental IPNV challenge, respectively. An IPNV carrier condition at low virus titre did not influence the mortality rates after secondary infections. Neither the ISAV infection nor the V. salmonicida infection in experimentally induced IPNV carriers resulted in mortalities different from those observed after challenge of IPNV-free fish. At higher IPNV titres in Atlantic salmon with acute IPN, the outcome of secondary infections was quite different from that observed in IPNV-free fish and in IPNV carriers. In 2 different experiments significantly more fish died when fish with acute IPN were infected with V salmonicida than when fish were infected with V salmonicida alone. Mortality also started earlier in the double-infected group than in the group challenged with V. salmonicida alone, 3 to 4 and 8 d after V salmonicida infection, respectively. Similar results were observed independent of whether mortalities due to IPN alone were registered in the experiments. When Atlantic salmon with acute IPN were infected with ISAV, significantly fewer fish died than when fish were infected with ISAV alone. The ongoing IPNV infection seemed to provide some protection against development of ISA.     

Evaluation of a water concentration method for the detection of infectious pancreatic necrosis virus in a fish hatchery

A virus-adsorption-elution (viradel) method using electropositive microporous filters (Virosorb 1-MDS) was used for the concentration and détéction of infectious pancreatic necrosis virus (IPNV) at a hatchery with a past history of IPNV infection. Samples of fish tissue and unconcentrated water were also examined for the presence of IPNV. Virus was isolated from three of the nine concentrated water samples taken from various locations at the hatchery. Virus was also isolated from pooled fish tissues, but not from unconcentrated water samples. Representative numbers of viruses isolated from water and fish tissues were examined for virus neutralization in the presence of anti-IPNV amiserum; resistance to acid, ether, and heat; and replication in the presence of 5-bromo-2'-deoxyuridine (BUDR). When tested by dot-immunobinding assay using a panel of monoclonal antibodies (MABs), the viral isolates were found to belong to the West Buxton serotype of IPNV.     

Effects of salmonid fish viruses on Mx gene expression and resistance to single or dual viral infections

We examined the ability of several fish viruses to induce protection against homologous or heterologous viruses in single or double infections, and assessed whether such protection is correlated with innate immunity or expression of the Mx gene. Monolayers of BF2 cells pre-treated with supernatants of brown trout (Salmo trutta L.) macrophage cultures that had been stimulated with either polyinosinic polycytidylic acid (poly I:C) or viruses, such as infectious pancreatic necrosis virus (IPNV), infectious haematopoietic necrosis virus (IHNV) or a mixture of the two, showed varying degrees of protection against viral infections. The virus showing the strongest induction was IPNV, and the antiviral activity against IHNV was also high: around 6 log(10) reduction of virus yield. Consequently, the IPNV-IHNV co-infection yield was also reduced by varying amounts. In vivo, the cumulative mortality observed in the IPNV-IHNV co-infected fish was always less than that in those with a single infection. Stimulation with poly I:C for 7 days significantly reduced cumulative mortality in single-infected fish, but not in the double-infected, in which the IPNV was the only virus isolated from moribund animals. By RT-PCR, Mx was expressed in all the organ samples tested (kidney, liver and spleen) from virus-stimulated fish at 1, 2 and 3 days. By qRT-PCR the extent and timing of Mx expression was shown to differ in the poly I:C and the single or dual viral infections. The highest increase in Mx expression (21.6-fold above basal levels) occurred (after 24 h) in fish infected with the IHNV, and expression remained high until day 7. Mx expression in fish infected with IPNV peaked later, at 2 days post infection, and also remained high until day 7. The dual infection with IPNV-IHNV induced high Mx expression on day 1, which peaked on day 2 and remained high until day 7. These results indicate that activation of the immune system could explain the interference and loss of IHNV in the IPNV-IHNV co-infections.     

Slow release cortisol implants result in impaired innate immune responses and higher infection prevalence following experimental challenge with infectious pancreatic necrosis virus in Atlantic salmon (Salmo salar) parr

Stress can affect the immune system and increase susceptibility to various diseases but knowledge of the underlying mechanisms is scarce. There is a complex interaction between the immune system and the endocrine system of vertebrates. In fish, cortisol is a key hormone regulating stress response and recent studies have also suggested that this hormone can affect the immune system, where cortisol is mainly regarded as an immunosuppressive factor. The aim of the present study was to examine the impact of chronically elevated levels of cortisol on the immune response and susceptibility to experimental infection with infectious pancreatic necrosis virus (IPNV). Further, the effect of IPNV challenge on circulating levels of cortisol was investigated. Atlantic salmon parr were implanted intraperitoneally with sustained-release implants of bovine of cortisol (50 μg cortisol g(-1) body weight in an implant based on vegetable lipids). Vehicle implants were used as control (sham-injected). At 45 days after implantation (DAI), fish were challenged with a low virulent isolate of IPNV (by immersion). Samples of plasma, liver and head kidney was taken from fish before and 24 h, 48 h, 7 days week and 21 days post infection (DPI). Cortisol level in plasma was measured using radioimmunoassay and gene expression in liver and head kidney was analyzed with real-time PCR (RT-PCR). Infection prevalence in infected fish was assessed by virus culture and RT-PCR of head kidney samples. Cortisol implantation compared with sham-implanted fish had increased levels of plasma cortisol at 45 DAI. The relative expression of Interferon alpha-1 (IFNα-1), Myxo virus-1 Mx, Heat-shock protein 70 (HSP70), Serum amyloid A (SAA), Glucocorticoid receptor (GR) and Heat-shock protein 90 (HSP90) tends to be down-regulated by cortisol implantation. There was a higher prevalence of fish with detectable levels of IPNV, as measured by cell culture and RT-PCR, in the cortisol-implanted group challenged with IPNV (0 = 0.0305) relative to the group that received a sham implantation. Further, cortisol seems to delay the induction of the antiviral IFNα-1 pathway and Mx mRNA expression. This study shows that elevated plasma cortisol level leads to an impaired innate immune response, and higher virus (IPNV) prevalence in Atlantic salmon parr.     

Isolation of salmon pancreas disease virus (SPDV) in cell culture and its ability to protect against infection by the 'wild-type' agent

A Scottish salmon pancreas disease virus (SPDV) has been isolated and its optimum growth conditions determined. Although several fish cell lines have been tested, successful culture was achieved only with CHSE-214 cells. Cytopathic effects were observed after 5 days. The highest virus titres, calculated by microtitration assay, were reached at 15 degrees C. After 7-9 days post-inoculation, CHSE-214 cell supernatants contained between 10(7)-10(5) TCID50 ml(-1) The cultured isolate is chloroform- and pH 3.0-sensitive, and virions are 50-60 nm in diameter. These characteristics are similar to the Irish SPDV isolates. The culture isolate induced typical pancreas disease (PD) lesions in experimentally infected Atlantic salmon and convalescent fish were resistant to experimental infection with PD-infective kidney homogenates obtained by serial in vivo passages from a PD-infected farmed salmon (termed wild-type SPDV). Furthermore, fish immunised with the inactivated cultured virus were protected against a cohabitation challenge with the wild-type virus. Immunised fish sera showed virus-neutralising activity before challenge (7 weeks post-immunisation) and from 3-6 weeks post-challenge, when sera from non-immunised fish did not neutralise the virus. At 6 weeks post-cohabitation challenge, previously immunised fish had neutralising titres of up to 1:65. Following intraperitoneal (i.p.) challenge, immunised fish showed neutralising titres as high as 1:226 at 8 weeks post-challenge. Non-immunised fish injected i.p. with the wild-type virus developed serum-neutralising activity against the cultured isolate when sampled 8 weeks after infection, confirming an antigenic relationship between the wild-type and cultured virus. The results demonstrate that the tissue culture-adapted isolate of SPDV could be successfully used to protect against challenge by the wild-type virus and could therefore have potential use as an inactivated vaccine against PD.     

鳜鱼病毒病原的检出及组织病理分析

经对患暴发性传染病的鳜组织及病变组织超微切片的电镜观察,在细胞质,核及间隙中都有病毒颗粒,对该病毒进行了分离提纯,这是一种直径约为２８０ｎｍ球形病毒,将粗提病毒液接种到培养细胞中,会引起细胞病变;病毒通过人工感染健康鳜,会引起鳜发病和死亡,对患病獗进行解剖和组织病理观察,表明病鳜的５种器官都发生了病变,研究结果表明分离的鳜病毒（ＳｉｎｉｐｅｒｃａｃｈｕａｔｓｉＶｉｒｕｓ,ＳＣＶ）是引起鳜流４行的病     

Dynamics of nontypical apoptotic morphological changes visualized by green fluorescent protein in living cells with infectious pancreatic necrosis virus infection

Morphologically, apoptotic cells are characterized by highly condensed membrane blebbing and formation of apoptotic bodies. Recently, we reported that apoptosis precedes necrosis in a fish cell line infected with infectious pancreatic necrosis virus (IPNV). In the present study, we tested the possibility that nontypical apoptosis is a component of IPNV-induced fish cell death. A variant type of green fluorescent protein (EGFP) was expressed in a fish cell line such that EGFP served as a protein marker for visualizing dynamic apoptotic cell morphological changes and for tracing membrane integrity changes during IPNV infection. Direct morphological changes were visualized by fluorescence microscopy by EGFP in living cells infected with IPNV. The nontypical apoptotic morphological change stage occurred during the pre-late stage (6 to 7 h postinfection). Nontypical apoptotic features, including highly condensed membrane blebbing, occurred during the middle apoptotic stage. At the pre-late apoptotic stage, membrane vesicles quickly formed, blebbed, and were finally pinched off from the cell membrane. At the same time, at this pre-late apoptotic stage, apoptotic cells formed unique small holes in their membranes that ranged from 0.39 to 0.78 micrometer according to examination by scanning electron microscopy and immunoelectron microscopy. Quantitation of the intra- and extracellular release of EGFP by CHSE-214-EGFP cells after IPNV infection was done by Western blotting and fluorometry. Membrane integrity was quickly lost during the late apoptotic stage (after 8 h postinfection), and morphological change and membrane integrity loss could be prevented and blocked by treatment with apoptosis inhibitors such as cycloheximide, genistein, and EDTA before IPNV infection. Together, these findings show the apoptotic features at the onset of pathology in host cells (early and middle apoptotic stages), followed secondarily by nontypical apoptosis (pre-late apoptotic stage) and then by postapoptotic necrosis (late apoptotic stage), of a fish cell line. Our results demonstrate that nontypical apoptosis is a component of IPNV-induced fish cell death.     

Isolation of different types of birnavirus from ayu Plecoglossus altivelis and amago salmon Oncorhynchus rhodurus cultured in the same geographic area

A birnavirus was recently isolated from cultured ayu Plecoglossus altivelis on Shikoku island, Japan. The diseased fish displayed vertebral or vertical curvature and mild haemorrhage around the brain. Cytopathic effects (CPE) of the virus, including cell roundness, filamentous change and cell lysis, were observed in CHSE-214, RTG-2 and RSBK-2 cells. The virus isolated from ayu, designated the AY-98 strain, was found to be antigenically related to the marine birnavirus (MABV) Y-6 strain that originated from yellowtail Seriola quinqueradiata. AY-98 had a bi-segmented RNA genome and the same nucleotide sequence in the 310 bp VP2/NS junction as MABV Y-6. At the same time that the ayu epizootics occurred, another birnavirus (AM-98) was isolated from amago salmon Oncorhynchus rhodurus which were cultured 66 km away from the ayu farm. AM-98 showed a similar CPE and had the same host cell ranges as AY-98. However, AM-98 was serologically similar to the VR-299 strain of infectious pancreatic necrosis virus (IPNV) and their nucleotide sequences in the VP2/NS junction region showed 98% homology without changes at the amino acid level. In this study, the ayu strain AY-98 was grouped into MABV, whereas the amago salmon strain AM-98 was grouped into IPNV. This indicates that the 2 birnaviruses originated from different sources in spite of the fact that the places where they were isolated are close to one another. The results in this paper show a new aspect of the traditional consensus that the same serogroup of birnavirus distribute in close geographic areas.     

Virus susceptibility of the fish cell line SAF-1 derived from gilt-head seabream

The recently reported SAF-1 cell line from fins of gilt-head seabream was evaluated for susceptibility to lymphocystis disease virus (LDV) and to several salmonid fish viruses, such as infectious haematopoietic necrosis virus (IHNV), viral haemorrhagic septicemia virus (VHSV) and several strains of infectious pancreatic necrosis virus (IPNV). LDV, VHSV and IHNV replicated well in the cultured fin cells as demonstrated by cell lysis and increases in viral titer. The potential use of this cell line to detect viruses from fish marine species is discussed.     

Establishment and characterization of the Epithelioma Papulosum Cyprini (EPC) cell line persistently infected with Infectious Pancreatic Necrosis Virus (IPNV), an aquabirnavirus

Infectious pancreatic necrosis virus (IPNV), a type species of aquabirnaviruses in the family Birnaviridae, is an etiological agent of infectious pancreatic necrosis and has been isolated from epizootics of cultured salmonids. In the present study, an epithelioma papulosum cyprini (EPC) cell line persistently infected with IPNV (PI-EPC) was experimentally established by subculturing EPC cells surviving IPNV infection, and was characterized. PI-EPC cells were morphologically indistinguishable from EPC, but continued to grow and yield IPNV. PI-EPC cells showed no cytopathic effect due to IPNV inoculation, and susceptibility of PI-EPC cells against heterologous viruses was not different from that of EPC cells. Only one cell of 10^3.5 PI-EPC cells produced IPNV at approximately 10^0.5 50% tissue culture infectious dose (TCID50)/cell/day, which was approximately 1,000 times lower than that of normal EPC cells. PI-EPC cells that did not yield IPNV (N-PI-EPC) were screened. The IPNV genome was detected from both PI-EPC and N-PI-EPC cells, and the IPNV VP2 structural protein was detected from both cell lines, but no other IPNV proteins were observed by Western blot analysis with anti-IPNV serum. Thus, multiplication of IPNV in PI-EPC cells was regulated by some host cell factors, except interferon.     

Pathogenicity and distribution of avian nephritis virus (G-4260 strain) in inoculated laying hens

Specific-pathogen-free laying hens were inoculated intravenously with the G-4260 strain of avian nephritis virus (ANV). The distribution of the virus in organs, histological changes in main organs, the condition of laying, and egg transmission of the virus were examined in them. Over an experimental period of 27 days, no clinical sings were observed. In a chronological study on the distribution of the virus in organs, the virus was recovered from liver, kidney, jejunum, and rectum for 6 days postinoculation (PI). The virus titer in organ emulsion was the highest in the jejunum of all the main organs. The virus was recovered from the kidney for 8 days PI, although it was not so high in this organ. It was not recovered from the ovary or oviduct. Fluorescent antigens were not observed at all in any material. In a pathological examination, some local inflammatory changes were observed only in the kidney. There were no significant changes in the ovary, oviduct, or any other organ. Antibody appeared 10 days PI and was detectable even 27 days PI, although it was not so high in titer. There was no significant difference in the rate of egg-production between the infected and the sham inoculated groups. No virus was isolated from 111 fertile eggs laid by infected hens over a period from 2 to 27 days PI.     

Viral susceptibility of newly established cell lines from the Hawaiian monk seal Monachus schauinslandi

Ten of 11 cell lines, recently established from the snout (MS-SN), periorbital soft tissue (MS-EY), liver (MS-LV), kidney (MS-KD), lung (MS-LG), spleen (MS-SP), heart (MS-HT), thyroid (MS-TY), brain (MS-BR) and urinary bladder (MS-UB) of a juvenile Hawaiian monk seal Monachus schauinslandi, were evaluated in vitro for their susceptibility to 5 mammalian viruses: herpes simplex virus type 1 (HSV-1), vesicular stomatitis virus (VSV), reovirus type 3 (Reo-3), poliovirus type 1 (Polio-1) and vaccinia virus (Vac); 5 fish viruses: channel catfish herpesvirus (CCV), infectious hematopoietic necrosis virus (IHNV), infectious pancreatic necrosis virus (IPNV), fish rhabdovirus carpio (RC) and viral hemorrhagic septicemia virus (VHSV); and 2 marine mammal morbilliviruses: phocine distemper virus (PDV) and dolphin distemper virus (DMV). Four well-established continuous cell-lines of nonhuman primate (Vero) and fish (EPC, CHSE-214 and BB) origin served as controls to standardize the virus infectivity assays. Virus yields were quantified as 50% tissue culture infectious dose (TCID50) ml(-1) on Day 7 post-inoculation. Results of the viral challenge assays revealed that the monk seal cell lines shared a similar pattern of susceptibility to the mammalian viruses. Despite their different tissue origins, all monk seal cells were sensitive to HSV-1, Vac, VSV and Reo-3, but were refractory to Polio-1. A characteristic viral-induced cytopathic effect was noted with VSV and Reo-3, and distinct plaques were observed for HSV-1 and Vac. Monk seal cell lines were also susceptible to PDV and DMV, 2 morbilliviruses isolated from seals and dolphins, respectively. By contrast, these cell lines were generally resistant to VHSV, IHNV and IPNV, with varying susceptibility to RC and CCV. The wide range of viral susceptibility of these monk seal cell lines suggests their potential value in studying viruses of monk seals and other marine mammals.     

Monitoring viral-induced cell death using electric cell-substrate impedance sensing

Using an electrical measurement known as electric cell-substrate impedance sensing (ECIS), we have recorded the dynamics of viral infections in cell culture. With this technique, cells are cultured on small gold electrodes where the measured impedance mirrors changes in attachment and morphology of cultured cells. As the cells attach and spread on the electrode, the measured impedance increases until the electrode is completely covered. Viral infection inducing cytopathic effect results in dramatic impedance changes, which are mainly due to cell death. In the current study, two different fish cell lines have been used: chinook salmonid embryonic (CHSE-214) cells infected with infectious pancreatic necrosis virus (IPNV) and epithelioma papulosum cyprini (EPC) carp cells infected with infectious hematopoeitic necrosis virus (IHNV). The impedance changes caused by cell response to virus are easily measured and converted to resistance and capacitance. An approximate linear correlation between log of viral titer and time of cell death was determined.     

Experimental Transmission of Infectious Pancreatic Necrosis Virus from the Blue Mussel,Mytilus edulis,to Cohabitating Atlantic Salmon (Salmo salar) Smolts

Integrated multitrophic aquaculture (IMTA) reduces the environmental impacts of commercial aquaculture systems by combining the cultivation of fed species with extractive species. Shellfish play a critical role in IMTA systems by filter-feeding particulate-bound organic nutrients. As bioaccumulating organisms, shellfish may also increase disease risk on farms by serving as reservoirs for important finfish pathogens such as infectious pancreatic necrosis virus (IPNV). The ability of the blue mussel (Mytilus edulis) to bioaccumulate and transmit IPNV to naive Atlantic salmon (Salmo salar) smolts was investigated. To determine the ability of mussels to filter and accumulate viable IPNV, mussels were held in water containing log 4.6 50% tissue culture infective dose(s) (TCID₅₀) of the West Buxton strain of IPNV ml⁻¹. Viable IPNV was detected in the digestive glands (DGs) of IPNV-exposed mussels as early as 2 h postexposure. The viral load in mussel DG tissue significantly increased with time and reached log 5.35 ± 0.25 TCID₅₀ g of DG tissue⁻¹ after 120 h of exposure. IPNV titers never reached levels that were significantly greater than that in the water. Viable IPNV was detected in mussel feces out to 7 days postdepuration, and the virus persisted in DG tissues for at least 18 days of depuration. To determine whether IPNV can be transmitted from mussels to Atlantic salmon, IPNV-exposed mussels were cohabitated with naive Atlantic salmon smolts. Transmission of IPNV did occur from mussels to smolts at a low frequency. The results demonstrate that a nonenveloped virus, such as IPNV, can accumulate in mussels and be transferred to naive fish.     

Notices

A virus isolated from the lung of an aborted Hereford fetus was shown to possess the physical and chemical properties of the herpesvirus group. The virus designated bovine herpesvirus (BH-1247) was isolated in cultures of Madin-Darby bovine kidney (MDBK) and primary bovine embryonic kidney (BEK) cells. Electron microscopic studies revealed typical forms of virus particles of the herpesvirus group. The virus was sensitive to chloroform and virus replication was inhibited by the addition of 5-iododeoxyuridine into the cell culture medium. The characteristic features of the cytopathic changes were syncytial formations with intranuclear inclusion bodies. Discrete plaques were formed in MDBK cell cultures overlaid with agar. Virus growth studies in BEK cells revealed infectious virus to be cell associated and replicated at low titer. By serum neutralization tests the virus was shown to be distinct from bovine herpesviruses; infectious bovine rhinotracheitis, DN-599, Movar 33/63, bovine mammallitis, malignant catarrhal fever and feline viral rhinotracheitis, equine herpesvirus I and pseudorabies. The isolate was nonpathogenic to mice inoculated subcutaneously, intracerebrally and intraperitoneally. Virus replication was not demonstrated when inoculated on the chorioallantoic membrane of embryonated eggs.