The First Isolation in North America of Infectious,Hematopoietic Necrosis Virus (IHNV) and Viral Hemorrhagic Septicemia Virus (VHSV) in Coho Salmon from the Same Watershed

Abstract

In November 1989, infectious hematopoietic necrosis virus (IHNV) was found for the first time in the Soleduck River at the Washington Department of Fisheries Soleduck Hatchery. The virus was isolated from ovarian fluid and kidney-spleen tissue pools from chinook salmon Oncorhynchus tshawytscha and ovarian fluid pools from coho salmon O. kisutch returning to the Soleduck Hatchery. The virus was identified as IHNV by neutralization assays. In December 1989, the virus causing viral hemorrhagic septicemia (VHSV) was found in ovarian fluid and milt pools from wild coho salmon obtained from the Soleduck and Bogachiel rivers and held at the Soleduck Hatchery. The virus was identified as VHSV by neutralization and immunoblot assays. These findings and their implications for routine broodstock sampling are discussed.     

Differing Resistance of Atlantic Salmon Strains and Rainbow Trout to Gyrodactylus salaris Infection

Abstract

Laboratory studies were conducted on the susceptibility of different strains of Atlantic salmon Salmo salar and rainbow trout Oncorhynchus mykiss to infection with the monogenean Gyrodactylus salaris. This parasite, probably originating from the Baltic Sea region, is known to minimally affect Neva River (Baltic Sea) Atlantic salmon. However, following its introduction into Norway, G. salaris has caused severe mortality and morbidity among Norwegian Atlantic salmon, which are considered a highly susceptible strain. The cohabitation experiment included one stock of rainbow trout and four different strains of Atlantic salmon from the Baltic Sea region (Mörrum River, Sweden), Europe (Skjern River, Denmark; Conon River, Scotland), and North America (Bristol Cove River, Canada). Fish were exposed to a Norwegian strain of G. salaris, and parasite population development and distribution were monitored for 7 weeks. Rainbow trout exhibited low susceptibility to G. salaris infection, whereas Conon River and Skjern River Atlantic salmon were highly susceptible and exhibited high mortality rates. Mörrum River Atlantic salmon exhibited intermediate susceptibility and low mortality. Bristol Cove River Atlantic salmon harbored relatively low parasite numbers, but fish mortality was high. Our experiment showed that the Danish Skjern River strain of Atlantic salmon is highly susceptible to G. salaris infection, further supporting the hypothesis that Atlantic Ocean strains are more susceptible to G. salaris infection than are Baltic strains.     

Primary Isolation and Characterization of Tenacibaculum maritimum from Chilean Atlantic Salmon Mortalities Associated with a Pseudochattonella spp. Algal Bloom

Abstract

This study presents the first isolation of Tenacibaculum maritimum from farmed Atlantic Salmon Salmo salar in Chile. The isolate, designated T. maritimum Ch-2402, was isolated from gills of Atlantic Salmon at a farm located in region X, Los Lagos, Chile, during the harmful algal bloom caused by Pseudochattonella spp. in February 2016. The algal bloom is reported to have caused 40,000 metric tons of mortality in this salmon farming area. The bacterium T. maritimum, which causes tenacibaculosis, is recognized as an important pathogen of marine fish worldwide. Genetic, phylogenetic, and phenotypic characterizations were used to describe the T. maritimum Ch-2402 isolate. The isolate was similar to the type strain of T. maritimum but was genetically unique. Tenacibaculum dicentrarchi isolates were also recovered during sampling from the same farm. Based on the fact that T. maritimum has been shown to cause disease in Atlantic Salmon in other regions, the presence of this bacterium poses a potential risk of disease to fish in the Chilean aquaculture industry.

Received November 6, 2016; accepted May 29, 2017 Published online July 26, 2017     

Infectious Hematopoietic Necrosis Virus (IHNV) in Coho Salmon

Abstract

Infectious hematopoietic necrosis virus (IHNV) causes important losses of chinook salmon Oncorhynchus tshawytscha, sockeye salmon Oncorhynchus nerka, and rainbow trout and steelhead Oncorhynchus mykiss on the west coast of North America. Although coho salmon Oncorhynchus kisutch are considered resistant to IHNV infection, the virus was detected in numerous adult coho salmon returning to Trinity River Hatchery, California, in 1985 and 1986. The virus was isolated from internal organs and ovarian fluids of these fish. Antigenic and structural polypeptides of the viruses were identical in adult coho and chinook salmon collected at the same location. Chinook salmon and rainbow trout alevins exhibited high degrees of susceptibility to IHNV obtained from adult coho and chinook salmon. Coho salmon alevins were resistant to both virus isolants.     

The persistence of infectious pancreatic necrosis virus in Atlantic salmon

Atlantic salmon leucocytes from Infectious Pancreatic Necrosis Virus (IPNV) carriers showed a suppressed response to phytohemagglutinin stimulation compared with uninfected controls. A significant degree of inhibition of DNA synthesis was observed using 3H-thymidine incorporation. IPNV was isolated from 41% of the stimulated leucocyte cultures supernatants, while only 6% of the unstimulated cultures were found to be positive.     

Production of Ceratonova shasta Myxospores from Salmon Carcasses: Carcass Removal Is Not a Viable Management Option

Severe infection by the endemic myxozoan parasite, Ceratonova (synonym, Ceratomyxa) shasta, has been associated with declines in and impaired recovery efforts of populations of fall-run Chinook Salmon Oncorhynchus tshawytscha in the Klamath River, California. The parasite has a complex life cycle involving a polychaete worm host as well as a salmon host. Myxospore transmission of this parasite, from salmon to polychaete, is a life cycle step during which there is a potential for applied disease management. A 3-year data set on prevalence, intensity, and spore characteristics of C. shasta myxospores was obtained from adult Chinook Salmon carcasses surveyed in the main stem of the Klamath River and three of its tributaries, Bogus Creek and the Shasta and Trinity rivers. Annual prevalence of myxospore detection in salmon intestines ranged from 22% to 52%, and spore concentration values per intestinal scraping ranged from 3.94 × 10² to 1.47 × 10⁷ spores. A prevalence of 7.3% of all carcasses examined produced >5.0 × 10⁵ spores, and these carcasses with “high” spore counts accounted for 76–95% of the total spores in a given spawning season. Molecular analysis of visually negative carcasses showed that 45–87% of these samples had parasite DNA, indicating they contained either low spore numbers or presporogonic stages of the parasite. Myxospores were rarely found in carcasses of freshly spawned adults but were common in decomposed carcasses of both sexes. The date of collection or age (based indirectly on FL) did not influence detection. The longer prespawn residence time for spring-run Chinook Salmon compared with that for fall-run Chinook Salmon in the Trinity River was associated with higher spore loads. The dye exclusion method for assessing spore viability in fresh smears indicated an inverse relationship in spore integrity and initial spore concentration. A carcass-removal pilot project in Bogus Creek for 6 weeks in the fall of 2008 (907 carcasses removed) and 2009 (1,799 carcasses removed) failed to measurably influence the DNA quantity of C. shasta in targeted waters. Combined with the high numbers of carcasses that contributed myxospores, we therefore deemed that this labor-intensive approach is not a viable management option to reduce the infectivity of C. shasta in Chinook Salmon in the Klamath River.

Received January 23, 2015; accepted September 28, 2015     

Genotyping of Infectious Pancreatic Necrosis Virus Isolates from Mexico State

Abstract

The infectious pancreatic necrosis virus (IPNV; genus Aquabirnavirus) affects salmon and trout, causing high mortality in first-feeding fry. The classification of this virus includes nine serotypes and seven genogroups. In Mexico, two different isolates were identified in 2000 and 2008, respectively. Both isolates were classified into genogroup I according to the RNA genome of this virus. As Mexico is importing rainbow trout Oncorhynchus mykiss eggs from different countries, the aim of this study was to genotype IPNV isolates obtained from four rainbow trout producer regions within the state of Mexico. We utilized a fragment of the VP2* (outer capsid protein) gene sequence of Mexican IPNV isolates as a molecular marker to determine the genogroup to which they belong. Although all Mexican IPNV isolates were grouped into genogroup I, we identified genetic diversity among these isolates, and 14 unique nucleotide sequence types were associated with the four producer regions in Mexico State.

Received December 21, 2010; accepted July 27, 2011     

Survey of Pathogens in Hatchery Chinook Salmon with Different Out-Migration Histories through the Snake and Columbia Rivers

The operation of the Federal Columbia River Power System (FCRPS) has negatively affected threatened and endangered salmonid populations in the Pacific Northwest. Barging Snake River spring Chinook salmon Oncorhynchus tshawytscha through the FCRPS is one effort to mitigate the effect of the hydrosystem on juvenile salmon out-migration. However, little is known about the occurrence and transmission of infectious agents in barged juvenile salmon relative to juvenile salmon that remain in-river to navigate to the ocean. We conducted a survey of hatchery-reared spring Chinook salmon at various points along their out-migration path as they left their natal hatcheries and either migrated in-river or were barged through the FCRPS. Salmon kidneys were screened by polymerase chain reaction for nine pathogens and one family of water molds. Eight pathogens were detected; the most prevalent were Renibacterium salmoninarum and infectious hematopoietic necrosis virus. Species in the family Saprolegniaceae were also commonly detected. Pathogen prevalence was significantly greater in fish that were barged through the FCRPS than in fish left to out-migrate in-river. These results suggest that the transmission of infectious agents to susceptible juvenile salmon occurs during the barging process. Therefore, management activities that reduce pathogen exposure during barging may increase the survival of juvenile Chinook salmon after they are released.

Received May 27, 2010; accepted January 17, 2011     

Inoculation of infectious pancreatic necrosis virus serotype Sp did not cause pancreas disease in Atlantic salmon (Salmo salar L.).

Atlantic salmon were selected from a fish farm with no previous record of pancreas disease (PD) or infectious pancreatic necrosis virus (IPNV) infection. Groups of fish were inoculated with either IPNV (strain Sp) from cell culture, organ material from fish with PD or control material as phosphate-buffered saline (PBS). Virological, histological and immunohistochemical examinations were carried out throughout the experiment. None of the fish died or showed clinical symptoms of PD. Histological examination revealed no pathological changes, and immunohistochemical studies were negative. Virus was isolated only sporadically from the group inoculated with organ material, whereas it was isolated consistently from the group inoculated with virus propagated in cell culture, as well as from in-contact control fish after the first week. In a latent carrier test, changes were entirely lacking in the first mentioned group, and were only slight in the last mentioned group. The data suggest that PD is not a transmissible disease, and that IPNV isolated from a PD outbreak does not play any part in the etiology of this disease.     

Occurrence and Distribution of Aeromonas salmonicida Infections on Swedish Fish Farms, 1951–1987

Abstract

The occurrence of typical and atypical Aeromonas salmonicida infections on Swedish fish farms was surveyed, and the methods for controlling the disease are described here. This review is based on data for a period of 36 years from fish health control studies conducted by the National Veterinary Institute and the Salmon Research Institute. The epizootiology and routes of transmission of the pathogens are described, and policy and methods for disease control are discussed. The distribution of atypical A. salmonicida infections was entirely different from that of typical infections. There was also circumstantial evidence of different routes of transmission for typical and atypical A. salmonicida infections. Although subclinical carriers of atypical A. salmonicida occurred among Atlantic salmon Salmo salar and rainbow trout Oncorhynchus mykiss on fish farms, acute disease outbreaks were most frequent among Arctic char Salvelinus alpinus and brown trout Salmo trutta. On the other hand, acute outbreaks of typical A. salmonicida infections were frequent among Atlantic salmon and rainbow trout.     

Detection of infectious pancreatic necrosis virus (IPNV) RNA by hybridization with an oligonucleotide DNA probe

Marine farming of Atlantic salmon (Salmo salar) is growing rapidly in Norway, and fish diseases have now become one of the biggest obstacles for this new industry. Infectious pancreatic necrosis virus (IPNV) is commonly found in fish from Norwegian sea farms. Diagnosis of IPNV infection is, at present, mainly based on virus isolation in cell cultures and identification by neutralization tests (NT). A DNA-RNA hybridization assay was developed using a 24 base DNA oligonucleotide probe. This is homologous to a part of the nucleotide sequence of the IPNV genome coding for a protease. RNA extracted from IPNV and harvest from infected cell cultures were fixed to nylon filters and hybridized with the 32P end-labelled probe. The results showed that the probe specifically identified IPNV from these two materials, both for the three different virus strains (Ab, Sp and VR-299) used, and for several different field isolates. It did not hybridize with reoviruses or non-infected cell cultures used as controls. These results indicate that the probe is not serotype specific, and furthermore that RNA extraction is not required before hybridization. This method may be a useful alternative to NT for routine identification of IPNV, particularly when non-radioactive labelling of the probe is introduced.     

Ichthyophoniasis: An Emerging Disease of Chinook Salmon in the Yukon River

Abstract

Before 1985, Ichthyophonus was unreported among Pacific salmon Oncorhynchus spp. from the Yukon River; now it infects more than 40% of returning adult Chinook salmon O. tshawytscha. Overall infection prevalence reached about 45% in the Yukon River and about 30% in the Tanana River between 1999 and 2003. Mean infection prevalence was greater in females than males in the main-stem Yukon River during each of the 5 years of the study, but the infection prevalence in males increased each year until the difference was no longer significant. Clinical signs of ichthyophoniasis (presence of visible punctate white lesions in internal organs) were least at the mouth of the Yukon River (～10%) but increased to 29% when fish reached the middle Yukon River and was 22% at the upper Tanana River. However, clinical signs increased each year from 7% in 1999 to 27% in 2003 at the mouth of the river. As fish approached the upper reaches of the Yukon River (Canada) and the spawning areas of the Chena and Salcha rivers (Alaska), infection prevalence dropped significantly to less than 15% in females on the Yukon River and less than 10% for both sexes in the Chena and Salcha rivers, presumably because of mortality among infected prespawn fish. Age was not a factor in infection prevalence, nor was the position of fish within the run. The source of infection was not determined, but Ichthyophonus was not found in 400 Pacific herring Clupea pallasi from the Bering Sea or in 120 outmigrating juvenile Chinook salmon from two drainages in Alaska and Canada. Freshwater burbot Lota lota from the middle Yukon River were subclinically infected with Ichthyophonus, but the origin and relationship of this agent to the Chinook salmon isolate is unknown.     

Antigen dose and humoral immune response correspond with protection for inactivated infectious pancreatic necrosis virus vaccines in Atlantic salmon (Salmo salar L)

An enduring challenge in the vaccinology of infectious pancreatic necrosis virus (IPNV) is the lack of correlation between neutralizing antibodies and protection against mortality. To better understand the immunological basis of vaccine protection, an efficacy trial including Atlantic salmon (Salmo salar L.) vaccinated with a high antigen (HiAg) or low antigen (LoAg) dose vaccine was carried out in a cohabitation challenge model using the highly virulent Norwegian Sp strain NVI015. To pinpoint the immunological basis of vaccine protection, pathogenic mechanisms of IPNV were unraveled in control fish while obtaining feedback on mechanisms of protection in the vaccinated fish. During the incubation period, infection rates were highest in control fish, followed by the LoAg group with the lowest infections being in the HiAg group. Although both the liver and pancreas are target organs prone to tissue damage, infection in the liver was delayed until acute infection in most fish. A correlate of pathology determined as the cutoff threshold of viral copy numbers linked to tissue damage in target organs was estimated at ≥ 10^7.0, which corresponded with an increase in mortality. The kinetics of IFNα and Mx expression suggests that these genes can be used as biomarkers of IPNV infection progression. Mechanisms of vaccine protection involved reducing infection rates, preventing infection of the liver and reducing virus replication in target organs to levels below the correlate of pathology. Overall, the study shows that antigen dose corresponds with vaccine efficacy and that antibody levels can be used as a signature of protective immunity against pathological disease and mortality.     

Bacterial Analysis of Fertilized Eggs of Atlantic Salmon from the Penobscot,Naraguagus, and Machias Rivers,Maine

Serious losses have occurred at the U.S. Fish and Wildlife Service, Craig Brook National Fish Hatchery, East Orland, Maine, among eggs that were taken from Atlantic Salmon Salmo salar, which were held as captive broodfish during their returns to the Penobscot River, Naraguagus River, and Machias River to spawn. Bacterial isolations were attempted from external surfaces and the internal contents of individual eggs. Externally and in all cases, Pseudomonas fluorescens was the predominant bacterium associated with the surface of all eggs. These bacteria were resistant to a surface treatment of 1,667 ppm formalin for 15 min and, therefore, the monoclonal nature of P. fluorescens on egg surfaces was considered to result from its ability to resist the germicidal activity of formalin administered for antifungal treatments. Flavobacterium psychrophilum, the cause of bacterial coldwater disease, was isolated from the interior of 23.6, 18.1, and 29.2% of the dead Atlantic Salmon eggs from Penobscot River egg lots A-98, A-100, and A-101, respectively, and concentrations of this pathogen ranged from 1.0 × 10³ to >5 × 10⁸ CFU per gram of dead egg. Flavobacterium psychrophilum was also isolated from 8.3, 26.7, and 10.0% of the dead eggs from Naraguagus River egg lots N-158, N-161, and N-163, respectively, in which concentrations of this organism ranged from 1.0 × 10³ to 7.5 × 10⁸ CFU per gram of egg. This bacterium was also isolated from within 18.3% and 3.3% of the dead eggs from Machias River egg lots M-128 and M-142, respectively, and its concentrations ranged from 1.0 × 10³ to 1.5 × 10⁸ CFU per gram of egg. The finding of F. psychrophilum from within these eggs is indicative of this pathogen's widespread and persistent prevalence in Atlantic Salmon in New England.

Received December 19, 2014; accepted May 4, 2015     

Infectious Salmon Anemia (ISA) Virus: Infectivity in Seawater under Different Physical Conditions

Abstract

Infectious salmon anemia (ISA) virus (genus Isavirus, family Orthomyxoviridae), present in all major salmon producing countries, is the causative agent for a serious and commercially important disease affecting Atlantic Salmon Salmo salar. Nearly all ISA outbreaks occur in the marine production phase and knowledge about survival time for ISA virions in seawater is crucial for an adequate strategy to combat the disease. To acquire knowledge about this important factor, a study of ISA virus exposed to four different physical conditions was carried out. The virions’ survival was tested in sterile seawater, sterile seawater with normal ultraviolet light radiation (UVR), natural seawater, and natural seawater with UVR. During the 72-h experiment both presence of ISA virus RNA and the infectivity of ISA virions were monitored. The result of this study showed that the infectivity of ISA virions is lost within 3 h of exposure to natural seawater or sterile seawater with UVR. However, it was possible to detect ISA virus RNA throughout the experimental period. This indicates that the effect of both UVR and biological activity of natural seawater limits the survival time of ISA virions under normal conditions. The survival time of ISA virions in sterile seawater was less than 24 h. Based on the available literature and the present study it is not very likely that passive horizontal transmission in seawater over long distances can occur. This is due to the following factors: (1) the effect of UVR and biological activity on ISA virions infectivity found in the present study, (2) the speed and dilution effect in seawater currents in salmon farming areas, (3) the temperature during the major outbreak periods, and (4) the need for an infective dose of ISA virions to reach naive Atlantic Salmon.

Received August 22, 2013; accepted November 7, 2013     

Communications: The Occurrence of Myxobolus neurobius (Myxosporea) in Wild Young Atlantic Salmon and Arctic Char in Newfoundland

Abstract

Spores identified as Myxobolus neurobius were found in head homogenates of both wild young Atlantic salmon Salmo salar and Arctic char Salvelinus alpinus. Formalin-preserved spores from Atlantic salmon measured an average of 12.9 × 9.4 μm and contained two polar capsules that measured 6.4 × 2.9 μm. The spores from the Arctic char appeared morphologically identical by bright-field microscopy to those from the Atlantic salmon except they measured 13.7 × 9.6 μm and had polar capsules that measured 6.9 × 3.4 μm. Myxobolus neurobius has not previously been reported from Atlantic salmon or Arctic char.     

The Impact of Egg Ozonation on Hatching Success,Larval Growth,and Survival of Atlantic Cod,Atlantic Salmon,and Rainbow Trout

The direct exposure of fish eggs to ozonated water has generated interest as a means of ensuring pathogen-free eggs without the use of harsh chemicals. However, there are numerous knowledge gaps, including safe contact times, exposure levels, and potential long-term effects on aquaculture species in both freshwater and seawater. The effect of different ozone (O₃) doses (0.5–1.0, 1.5–2.0, and 2.5–3.0 mg of O₃/L for 90 s) on recently fertilized eggs of Atlantic Cod Gadus morhua and eyed eggs of Atlantic Salmon Salmo salar and Rainbow Trout Oncorhynchus mykiss was evaluated in comparison with the effects of two commercial disinfectants: Perosan (0.004 mg/L) and Ovadine (100 mg/L). The impact of ozone application was evaluated based on hatching success, larval nucleic acid concentration, larval growth, and survival. Overall, results indicated that ozonation of Atlantic Cod eggs at a dose less than 3.0 mg/L for 90 s produced no negative effect on the larvae up to 30 d posthatch. Furthermore, ozonation of Atlantic Salmon and Rainbow Trout eggs generated no negative effect on the larvae, based on monitoring until 85% yolk sac re-absorption (16 d posthatch).

Received May 6, 2014; accepted October 24, 2014