Early Kinetics of Infectious Hematopoietic Necrosis Virus (IHNV) Infection in Rainbow Trout

Abstract

A series of experiments was carried out with infectious hematopoietic necrosis virus (IHNV; 193-110 isolate) in rainbow trout Oncorhynchus mykiss (weight, ～1.2 g) to determine the duration of the patent period and the timing of onset of the infectious periods. We first attempted to transmit IHNV to recipient fish from infected rainbow trout 2–3 d after they had been exposed. No infection transfer occurred despite high titers (10^4.79 to 10^4.91 plaque-forming units 5–8 d postexposure (dpe). To determine the number of secondary cases produced by one infectious individual, we exposed approximately 50 rainbow trout (weight, ～1.5 g) in each of seven replicate tanks to a donor fish that had been infected with virus by bath exposure 3 d earlier. The prevalence of infection in recipient fish rose from 0.84% at 2 dpe to 7.9% at 6 dpe. Maximum incidence (22 cases) occurred between 2 and 4 dpe. No disease-specific mortalities occurred in recipient fish during the experiment. The titer of virus in both recipient and donor fish increased from 2 to 4 dpe. There was a positive correlation between the level of infection among donors and prevalence values among recipient fish (r ² = 0.60). The level of challenge by one infectious fish under the conditions provided was enough for infection transfer from sick cohabitant to susceptible fish but was not enough for initiation of a full-scale epizootic among recipients.     

Early Life Stage Survival and Susceptibility of Brook Trout,Coho Salmon,Rainbow Trout,and Their Reciprocal Hybrids to Infectious Hematopoietic Necrosis Virus

Abstract

Triploid (heat-shocked) and diploid groups of rainbow trout Oncorhynchus mykiss, brook trout Salvelinus fontinalis, coho salmon Oncorhynchus kisutch, and reciprocal hybrids were produced, monitored for early life stage survival, and evaluated for susceptibility to infectious hematopoietic necrosis virus (IHNV). The female rainbow trout × male brook trout triploid hybrids had significantly greater (P < 0.01) survival than the diploid hybrids of this cross. The heat-shocked hybrid group of the female rainbow trout × male coho salmon also exhibited significantly greater survival to the eyed egg stage of development than the untreated group of this hybrid. Studies of the susceptibility of treatment groups to a 1990 IHNV isolate from the Hagerman Valley were conducted by using a standardized immersion exposure procedure at one or two different mean body weights. The diploid brook trout and coho salmon and two triploid hybrids (female rainbow trout × male brook trout or male coho salmon) were significantly less (P < 0.05) susceptible to IHNV than the pure-species diploid and triploid rainbow trout groups.     

White Sturgeon as a Potential Vector of Infectious Hematopoietic Necrosis Virus

Abstract

Cell lines from white sturgeon Acipenser transmontanus were derived from peripheral blood cells, heart, and spleen. Incubated with infectious hematopoietic necrosis virus (IHNV) for 8 d at l5°C, these cell lines produced 0.7–53.2 plaque-forming units (PFU)/cell. Waterborne exposure of larval white sturgeons (60 d posthatch) to 10⁶ PFU/mL of IHNV resulted in 10% mortality 5–6 d postinfection, with virus concentrations consistently greater than 10⁵ PFU/g. A replicate group of larval white sturgeons that were sampled at different times post-IHNV exposure had no detectable virus at 24 h, but 72% of the fish had IHNV concentrations of 10²-10⁶ PFU/g when they were examined 2–9 d postinfection. Juvenile white sturgeons (mean weight, 35 g) immersed in or injected with IHNV exhibited no mortality, and virus was only detected immediately postexposure in just 25% of the fish tested. Juvenile white sturgeons fed either virus-free rainbow trout Oncorhynchus mykiss or dead IHNV-infected rainbow trout had no viable virus in their feces. Juvenile white sturgeons fed or exposed to IHNV failed to transmit the virus to cohabiting rainbow trout fry. These results suggest that IHNV can replicate in larval white sturgeons but presumably not in juveniles or adults. Virus neutralization activity was detected in serum from adult white sturgeons (4–6 years old) cultured with rainbow trout exposed to IHNV but not in white sturgeons kept in a pathogen-free environment and fed a manufactured diet. White sturgeon serum with IHNV-neutralizing activity was used to passively immunize rainbow trout, and it provided significant (P < 0.01) protection against IHNV challenge.     

Vaccination of Rainbow Trout against Infectious Hematopoietic Necrosis (IHN) by Using Attenuated Mutants Selected by Neutralizing Monoclonal Antibodies

Abstract

A neutralizing monoclonal antibody against infectious hematopoietic necrosis virus (IHNV) was used to select neutralization-resistant mutants from isolates of virus obtained from adult steelhead Oncorhynchus mykiss returning to the Round Butte Hatchery (RB mutants) on the Deschutes River in Oregon, USA, and from rainbow trout (nonanadromous O. mykiss) at a commercial hatchery in the Hagerman Valley of Idaho, USA (193-110 mutants). Two of the mutants, RB-1 and 193-110-4, were significantly (P < 0.001) attenuated compared with parental strains. Vaccination of rainbow trout by waterborne exposure to the mutants conferred solid protection against challenge with wild-type virus. In some trials, fish vaccinated with the RB-1 mutant at 50% tissue culture infectious doses (TCID50) of 1 × 10⁴–1 × 10⁵ TCID50/mL or with the 193-110-4 mutant at 1 × 10²–1 × 10³ TCID50/mL, held for 14 d, then challenged with the homologous wild-type strain at 1 × 10⁵ TCID50/mL showed relative percent survival of 95–100% (P < 0.005). There was no significant difference (P > 0.05) in protection among fish exposed to the RB-1 vaccine strain at a dose of 1 × 10⁵ TCID50/mL for periods of either 1, 12, or 24 h, held for 14 d, and then challenged with the wild-type RB isolate, although the 1-h exposure seemed to be somewhat less effective. Fish were vaccinated with the RB-1 strain at 1 × 10³–1 × 10⁵ TCID50/mL for 24 h then challenged after 1, 7, 14, or 21 d with the wild-type RB isolate. No significant (P > 0.1) protection was observed at 1 d postvaccination, but the relative percent survival increased progressively at each subsequent challenge period, becoming statistically significant by day 7 (P < 0.001) and beyond. These results suggested that resistance to challenge with wild-type virus resulted from development of IHNV-specific immunity and not from viral interference or interferon induction, and they reinforce the potential of an attenuated vaccine to control this important disease.     

Multiplication of Infectious Hematopoietic Necrosis Virus in Rainbow Trout following Immersion Infection: Organ Assay and Electron Microscopy

Abstract

Sequential spread of infectious hematopoietic necrosis virus (IHNV) to tissues of rainbow trout Oncorhynchus mykiss was examined following immersion infection with two different isolates of IHNV, a pathogenic strain and a nonpathogenic strain from rainbow trout. Virus strain 193–110 was highly pathogenic to 1-month-old rainbow trout and caused 100% mortality within 13 d, whereas strain RB-76 was much less virulent, causing 50% mortality by the 19th day. Virus titers of 1-month-old fingerling fish dying soon after infection were significantly higher than titers of those dying later. Assays of dissected tissues showed that gills of infected 2-month-old fingerlings contained virus as early as 16 and 20 h postinfection, with definite replication occurring at 48 h. The early presence of the virus in the gills followed shortly by appearance of the virus in the kidneys and spleen indicated that the virus spreads rapidly to the target organs. Virus was detected in many other organs at lower levels on the third day and increased to higher levels during the following days. Heart tissue had high titers later in the infection. When 4-month-old rainbow trout were infected with strain 193–110, the mortality was reduced and delayed, whereas those infected with strain RB-76 produced no mortality. Assays on the day of death of these older fingerlings infected with strain 193–110 revealed that fish dying soon after infection also had higher titers than those dying later. Electron microscopic examination offish organs showed the presence of typical IHNV particles budding off from various tissue cells of affected organs, including gill tissue. The destructive effect of the virus was particularly noticeable in the disarrangement of heart muscle organelles.     

Susceptibility of Selected Inland Salmonids to Experimentally Induced Infections with Myxobolus cerebralis,the Causative Agent of Whirling Disease

Abstract

Laboratory exposures to the infectious stages (triactinomyxons) of Myxobolus cerebralis demonstrated a range of susceptibility to whirling disease among four species of inland salmonids. Replicate groups of each species were exposed to two concentrations of triactinomyxons, a low dose (100–200 per fish) and a high dose (1,000–2,000 per fish). Exposed fish were evaluated for clinical signs, for severity of microscopic lesions at 35 d, 2 and 5 months, and for spore concentrations in the head cartilage at 5 months. A standard strain of rainbow trout Oncorhynchus mykiss matched for age served as a susceptible species control. Rainbow trout, westslope cutthroat trout O. clarki lewisi, Yellowstone cutthroat trout O. clarki bouvieri, and bull trout Salvelinus confluentus were susceptible to M. cerebralis infections. Clinical signs, including radical swimming (“whirling”) and black tails, were observed at 7 weeks postexposure among rainbow and cutthroat trout challenged at 3 weeks of age. Clinical signs were rare among bull trout exposed at an age of 4 weeks and absent among rainbow and cutthroat trout exposed at 3 months posthatch. Most rainbow, cutthroat, and bull trout were found to be infected when examined at 5 months postexposure. The most severe microscopic lesions among infected fish at 5 months postexposure were found among rainbow trout. Cutthroat trout had less severe lesions, bull trout had mild infections, and no evidence of infection was found among Arctic grayling Thymallus arcticus. Mean spore concentrations among infected fish correlated with the severity of microscopic lesion scores. Rainbow trout had mean concentrations of spores in head cartilage reaching 10⁶, whereas more resistant species such as bull trout had 10⁴ spores; no spores were found among Arctic grayling at 5 months postexposure.     

In Vitro Infection of Salmonid Epidermal Tissues by Infectious Hematopoietic Necrosis Virus and Viral Hemorrhagic Septicemia Virus

Abstract

The ability of two rhabdoviruses, infectious hematopoietic necrosis virus (IHNV) and viral hemorrhagic septicemia virus (VHSV), to infect fish skin was investigated by in vitro infection of excised tissues. Virus replication was determined by plaque assay of homogenized tissue extracts, and the virus antigen was detected by immunohistology of tissue sections. Gill, fin, and ventral abdominal skin tissues of rainbow trout Oncorhynchus mykiss that had been infected in vitro with a virulent strain of IHNV (193–110) produced substantial increases in virus titer within 24 h. Titers continued to increase up until day 3 of incubation; by this time, virus had increased 1,000-fold or more. This increase in IHNV titer occurred in epidermal tissues of fingerlings and of older fish. In another experiment, IHNV replicated in excised rainbow trout tissues whether the fish had been subject to prior infection with a virulent strain of IHNV (Western Regional Aquaculture Consortium isolate) or whether the fish had been infected previously with an attenuated strain of the virus (Nan Scott Lake, with 100 passes in culture). A virulent strain of VHSV (23/75) replicated effectively in excised gill tissues and epidermal tissues of rainbow trout and chinook salmon O. tshawytscha; however, the avirulent North American strain of VHSV (Makah) replicated poorly or not at all.     

Experimental induction of the carrier state in yearling brook trout: a model challenge protocol for IPNV immunization

Brook trout fry (Salvelinus fontinalis) were not protected from infectious pancreatic necrosis virus (IPNV) challenge by immersion vaccination with inactivated, purified virus at concentrations of 10(7) to 10(9) pfu/ml. Mortalities in vaccinated groups were higher than for the unvaccinated control group and appeared to be dose-dependent. A challenge protocol for adult brook trout was developed for future vaccine trials. A single intraperitoneal injection of virulent, purified virus was sufficient to make long-lasting carriers of 16 month-old trout. Fish underwent a transient viremia, identified by virus isolation from plasma and leucocytes. Feces were the most reliable samples for identification of IPNV carriers by non-sacrificial testing. Many fish in the remaining infected group were still carriers 12 and 27 weeks post-infection.     

Survivability of Infectious Hematopoietic Necrosis Virus in Fertilized Eggs of Masu and Chum Salmon

Abstract

The possibility of vertical transmission of infectious hematopoietic necrosis virus (IHNV) was studied with the eggs of masu (cherry) salmon Oncorhynchus masou and chum salmon O. keta. The surfaces of eggs and sperm were contaminated with IHNV (10^3.8-10^4.8 50% tissue culture infective dose [TCID50]/egg) and then the eggs were fertilized. Eggs just after fertilization and embryonated eggs also were infected by injection with IHNV (10^3.8 TCID50/egg) directly into the yolk. During incubation, eggs were held in running water at 10°C. Mortality of the eggs or hatched progeny was determined and isolation of IHNV on the surface or inside of the eggs was determined during the incubation period. No mortality occurred and no virus was detected in fertile eggs from contaminated gametes. For injected eggs, IHNV was not detected on the surface of masu and chum salmon eggs after 1 d of incubation. Infectivity of IHNV inside the eggs decreased gradually and could not be detected after 1 month of incubation. This rate of IHNV reduction in the fertilized egg was similar to that found in a mixture of IHNV and homogenized yolk contents. Several individual yolk components also showed anti-IHNV activity. When eyed eggs were injected with IHNV, the embryos of both masu and chum salmon became infected, and the concentration of virus increased rapidly and reached more than 10^6.5 TCID50/fish. The cumulative mortality of eggs injected at the eyed stage for both masu and chum salmon was 90%. The susceptibilities of hatched-out larvae of masu and chum salmon to IHNV were different; cumulative mortality was more than 90% in masu salmon and 20–30% in chum salmon artificially infected with the virus. We concluded that vertical transmission of IHNV is doubtful because the virus is apparently unable to survive in eggs before the eyed stage.     

DNA Vaccination Partially Protects Muskellunge against Viral Hemorrhagic Septicemia Virus (VHSV-IVb)

Abstract

A DNA vaccine containing the glycoprotein (G) gene of the North American viral hemorrhagic septicemia virus (VHSV) genotype IVb was developed to evaluate the immune response of fish following vaccination and evaluate its efficacy in protecting a susceptible species, the Muskellunge Esox masquinongy, against VHSV-IVb challenge. Seven weeks (539 degree-days) following vaccination with 10 μg of either pVHSivb-G or a control plasmid, Muskellunge were challenged by immersion with 10⁵ plaque-forming units (pfu)/mL of VHSV-IVb. Fish vaccinated with pVHSivb-G had a relative percent survival (RPS) of 45%. Vaccinated fish also had significantly lower mean viral titers in tissues (4.2 × 10² pfu/g) and viral prevalence (4%) than fish receiving the plasmid control vaccine (3.3 × 10⁵ pfu/g; 82%). Neutralizing antibodies were detected 28 d (308 degree-days) postchallenge (11 weeks postvaccination) in 100% of Muskellunge vaccinated with pVHSivb-G compared with only 12% of plasmid-control-vaccinated Muskellunge, suggesting robust induction of a secondary, adaptive immune response. In addition, pVHSivb-G–vaccinated Rainbow Trout Oncorhynchus mykiss challenged 7 d (100 degree-days) postvaccination with the heterologous novirhabdovirus, infectious hematopoietic necrosis virus (IHNV), experienced an RPS of 61%, compared to control fish, suggesting induction of an early and transient nonspecific antiviral immune response. This study provides an important starting point for VHSV-IVb vaccine development and useful information about the antiviral immune response elicited by DNA vaccination in a nondomesticated fish species.

Received May 1, 2016; accepted September 1, 2016     

Serological Evidence of Infectious Hematopoietic Necrosis in Rainbow Trout from a French Outbreak of Disease

Abstract

Following the detection of infectious hematopoietic necrosis virus (IHNV) in France in April 1987, a serological survey was conducted of the rainbow trout Oncorhynchus mykiss (formerly Salmo gairdneri) from an infected cultured stock previously known to be contaminated with viral hemorrhagic septicemia virus (VHSV) for 3 years. The work lasted from April to December 1987, at which time all the remaining fish were slaughtered. Serum samples were assayed by a plaque-reduction test and a simplified neutralization test that is more suitable for processing large numbers of serum samples. Such investigations revealed that IHNV neutralization by trout antibodies depended on trout complement, as did neutralization of VHSV. Incubation for 16 h at 4°C increased the sensitivity of the test compared to incubation for 1 h at 20°C. During the course of clinical IHN from April to June, young fish did not display any neutralizing activity, but in September, 29 of 50 of them exhibited significant anti-IHN neutralizing antibody titers ranging from 21 to over 160, and 18 of 46 of these same fingerlings did so in December. Similarly, fish that had undergone VHS infection in August began to develop anti-VHSV antibodies in December (5 of 50), demonstrating that one fish can harbor neutralizing antibodies to both IHNV and VHSV, and that these antibodies had required 14 weeks to appear under fish culture conditions at 10°C. As could be expected from seroneutralization tests, neutralizing antibodies to IHNV did not result in protection against VHS. Sera from 13 of 20 adult fish sampled in mid-June revealed neutralizing antibodies to IHNV, suggesting that they harbored the virus prior to the clinical infection that affected their progeny. Only two of the fish showed low anti-VHSV antibody titers. Similarly, neutralizing antibodies to IHNV were detected in 53 of 73 other adult fish sampled in late October, 10 months after they had spawned and 7 months after mortality had occurred among their progeny. Given the prevalence, level, and persistence of neutralizing antibody titers, the seroneutralization test would be worth investigating more thoroughly to define the conditions that could make it a reliable tool for checking the virus status of trout carriers.     

Infectious Pancreatic Necrosis Virus: Transmission from Infectious to Susceptible Rainbow Trout Fry

Abstract

Fry of rainbow trout Oncorhynchus mykiss were exposed to serotype VR-299 of infectious pancreatic necrosis virus (IPNV) by using a standardized immersion challenge. In concurrent experiments, fish were monitored for 11 d for excretion of IPNV or monitored for 9 d for excretion and transmission of IPNV to susceptible rainbow trout fry. Immersion-challenged fish began excreting virus within 2 d after challenge. The rate of IPNV excretion per fish increased steadily from about day 4 to day 8 and then decreased. Virus concentrations in tissues of immersion-challenged fish increased exponentially. Susceptible fish became infected with IPNV within 4 d after being introduced to immersion-challenged fish (e.g., 2 d after the challenged fish began excreting virus). By 9 d, 84% of the susceptible fish were infected with IPNV.     

Multiplication of Infectious Hematopoietic Necrosis Virus in Rainbow Trout following Immersion Infection: Whole-Body Assay and Immunohistochemistry

Abstract

The sites of replication of infectious hematopoietic necrosis virus (IHNV) in infected tissues were detected in fingerling rainbow trout Oncorhynchus mykiss by in situ histologic techniques following immersion infection. Virus antigens in tissues were detected by a neutralizing mouse monoclonal antibody and a one-step anti-mouse biotin-streptavidin conjugated to horseradish peroxidase. The efficiency of infection and virulence of the virus determined by mortality rates showed high virulence of the selected IHNV isolates, and viral replication in individual fish showed that virus content of the fish increased rapidly from the second day to the seventh day postinfection. The earliest viral lesions following infection were detected in the epidermis of the pectoral fins, opercula, and ventral surface of the body. Virus lesions became evident in kidneys on the third day. By the fifth day, when there was a significant increase in virus titer, foci of viral replication were detected in gill tissue and in the anterior internal tissues below the epidermis. Subsequently, extensive virus replication and tissue destruction were observed in the spleen, dorsal adipose tissues, ventricle, and pseudobranch. Replication in the liver, the muscularis layers of the digestive tract, and the general body musculature followed later. These infection experiments indicated that the epidermis and gills of fish constitute important sites of early IHNV replication.     

Biochemical and Antigenic Properties of the First Isolates of Infectious Hematopoietic Necrosis Virus from Salmonid Fish in Europe

Abstract

The first isolates of infectious hematopoietic necrosis virus (IHNV) recovered from rainbow trout Oncorhynchus mykiss (formerly Salmo gairdneri) in France and Italy were compared to six representative strains from North America by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) of virion polypeptides and neutralization by monoclonal antibodies (MAbs). All three IHNV isolates from Europe had similar polypeptide profiles when compared by SDS-PAGE. An analysis of the antigenic relatedness of the European isolates to representative strains from North America showed that they were clearly different from viruses obtained from salmonids in California. The RB/B5 MAb, which was developed against virus isolated from adult steelhead (anadromous rainbow trout) reared in central Oregon, neutralized all isolates examined. The 193–110/B4 MAb, developed against IHNV isolated from infected yearling rainbow trout in southern Idaho, neutralized all isolates tested except those from California. The SRCV/A4 MAb, developed against Sacramento River chinook virus (SRCV) isolated from adult spring chinook salmon O. tshawytscha in central California, was the least reactive, and strong neutralization was observed only with the SRCV strain of IHNV from California. However, partial reactivity of the virus isolates from France with the SRCV/A4 MAb distinguished them from the virus recovered from salmonids in Italy.     

Susceptibility of Four New Salmonid Cell Lines to Infectious Hematopoietic Necrosis Virus

Abstract

Four salmonid cell lines, CoE 45, CoE 115, CoE 345, and RBTE 45, were established from embryonic tissues of coho salmon Oncorhynchus kisutch and rainbow trout O. mykiss. In vitro challenges of the new lines were conducted with four isolates of infectious hematopoietic necrosis virus (IHNV). Two of the IHNV isolates used for the challenges were derived from infected tissues of rainbow trout, one was derived from chinook salmon O. tshawytscha, and the other isolate was derived from coho salmon. To standardize the virus challenges of the new cell lines, several established piscine cell lines (EPC, CHSE 214, CSE-119, RTH-149, RTG, and RTS) were challenged in the same way as the new lines. Each of the lines was challenged with virus at a single low multiplicity of infection (0.01 plaque-forming unit per cell). Virus yields were quantitated by plaque assay on epithelioma papulosum cyprini (EPC) cells on day 3. Results of the challenge experiments revealed different levels of production of virus for each isolate on the various cell lines. Overall, the new cell line derived from rainbow trout, RBTE 45, was quite susceptible to all viruses tested. The three cell lines newly derived from coho salmon embryo were not as resistant to the replication of IHNV as was the established coho salmon cell line, CSE-119. An established cell line, EPC, derived from an epithelial tumor of common carp Cyprinus carpio, remained the most susceptible to all four IHNV isolates tested.     

Fluorescent Antibody Test for the Rapid Diagnosis of Infectious Hematopoietic Necrosis

Abstract

A fluorescent antibody test (FAT) was developed for the rapid detection of infectious hematopoietic necrosis virus (IHNV). Both polyclonal and monoclonal antisera prepared against IHNV were evaluated. Test variables investigated included type of fixative, dilution rate of antibody reagents, staining time, and type of fluorescent conjugate that would be optimal for detection of IHNV. Specificity tests of the FAT indicated no cross-reactivity of the two antisera with other viruses or with cell lines of salmonid and nonsalmonid origin. All strains of IHNV tested, which included different electropherotypes, those isolated from selected salmonids at different life stages, and those from different geographic regions, reacted with both antisera. The FAT has been used for the detection of IHNV in blood smears and organ imprints from clinically infected juveniles, and in IHNV-infected cells in ovarian fluid from adult carriers. With this FAT, IHNV was detected after 48 h in cell lines inoculated with infected fish tissue. The test was equal in sensitivity to the plaque assay method and required less time to obtain a definitive diagnosis.     

Effects of Elevated Water Temperature and Immunosuppressant Treatment on Prevalence and Titer of Infectious Pancreatic Necrosis Virus in Naturally Infected Brook Trout

Abstract

Using fingerlings of brook trout Salvelinus fontinalis naturally infected with infectious pancreatic necrosis virus (IPNV), we demonstrated that elevated water temperature and treatment with the immunosuppressant triamcinolone acetonide (generic Kenalog®) significantly increases the titer of IPNV and probably also the prevalence of the infection. Stress-promoting treatments could potentially enhance the capability to detect various fish viral agents.     

Modulation of juvenile brook trout (Salvelinus fontinalis) cellular immune system after Aeromonas salmonicida challenge

In fish, the first line of defence against infectious microorganisms is based on non-specific cellular immune mechanisms (innate immunity). In this study, we measured the non-specific immune parameters (natural cytotoxic cells (NCC) activity, lymphoproliferation, percentage of phagocytosis and phagocytic activity) in brook trout (Salvelinus fontinalis) infected by a virulent strain of Aeromonas salmonicida. Eight days post-infection, the mortality of infected fish reached 70%. A transient immunostimulation of the NCC activity was noticed 24h post-infection, but there was no significant difference at 48 h. Then, infection of brook trout with A. salmonicida induced a biphasic immune response. At 24h post-infection, lymphoproliferation was drastically depressed but returned to control level at 96 h. A slight increase in the percentage of phagocytosis and the phagocytic activity was noticed throughout the experiment. Conversely the cell mortality was significantly higher in infected fish compared to control. The modulation of immunological parameters might reveal important clues on how innate immunity might protect fish from bacterial infections.     

Glycoprotein from Infectious Hematopoietic Necrosis Virus (IHNV) Induces Protective Immunity against Five IHNV Types

Abstract

Infectious hematopoietic necrosis virus (IHNV) is a fish pathogen that kills young salmon and trout. Outbreaks of the disease among hatchery-reared fish are a problem in the northwestern USA from northern California to Alaska. At least five biochemical types and several strains of differing host specificity of IHNV exist. Any vaccine developed to immunize fish must be able to elicit a response that will neutralize all strains of IHNV. This report shows that a single type of IHNV can induce a protective immune response in vivo to the five biochemical types of IHNV and indicates that, of the IHNV isolates examined, there is at least one common major neutralization epitope. Therefore, a vaccine developed against this common neutralization epitope will provide cross-protective immunity against these IHNV variants.     

Comparison of methods for detection of an infection with different isolates of infectious haematopoietic necrosis virus (IHNV)

The infectious haematopoietic necrosis (IHN) is beside the viral haemorrhagic septicemia (VHS) one of the viral fish diseases that have a considerable economic impact on German aquaculture. The measures actually in force are focused on control and spread prevention of the disease within the borders of the European Union (EU). The detection and confirmation of an outbreak is performed according to the pertinent EU legislation which allow the application of methods like the virus neutralisation test (VNT), the immunofluorescence test (IFT) and the enzyme-linked immunosorbent assay (ELISA). Besides the classic virological serology methods, further tests for the identification and confirmation of the fish pathogen like i.e. PCR and DNA probe techniques are recommended by the OIE. To compare diagnostic methods as ELISA, cell cultivation and RT-PCR, rainbow trout of the strain "Isle of Man" were infected with six IHNV strains. Samples were taken on day 7 (viraemia period) and at the day 28 of the trial. The ground organs were inoculated into EPC cells (Epithelioma papulosum Cyprini cells) and examined by ELISA as well as after RNA extraction by RT-PCR. Besides the determination of the isolate as well as the virulence for 20 g trout, significant differences in the demonstration of the viruses were observed. While the RT-PCR demonstrated to be the most sensitive method, antigen ELISA and virus cultivation results showed in dependence of the IHNV isolate that not all viruses were identifiable under the chosen experimental condition in the same manner.