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Prevention and control of viral diseases of salmonids

Three viral diseases of salmonids are of worldwide concern: infectious pancreatic necrosis (IPN) viral hemorrhagic septicemia (VHS), and infectious hematopoietic necrosis (IHN). Six principal approaches are being used to prevent or control these diseases: 1) preventing contact o the pathogen with the host, 2) environmental manipulation, 3) immunization, 4) chemotherapy, 5 selective breeding for disease resistance, and 6) reducing stress conditions which augment disease conditions. Preventing the introduction of a pathogen into a new stock of fish has been accomplished mainly by implementing stringent laws to prevent transport of infected fish into uninfected areas. Stocks of fish already infected are sometimes destroyed, and the hatchery is disinfected and restocked with fish free of specific pathogens. Environmental manipulation (elevated water temperature) has been successfully used to control IHN. Chemotherapeutics such as povidone-iodine for IPN and benzipyrene for IHN show promise of controlling mortalities; however, the practicality of using these drugs to eliminate the carrier fish has not been evaluated. Salmonids are capable of developing immune responses to viruses; however, development of effective vaccines, selective breeding for disease resistance, and identification of stress conditions which augment disease are still in the experimental phase.

Journal of the Fisheries Research Board of Canada

Scolecobasidium humicola , a fungal pathogen of fish

Scolecobasidium humicola , a previously undescribed fungal pathogen of fish was isolated from coho salmon ( Oncorhynchus kisutch ). In natural infections the kidney was the organ most affected. The disease was difficult to transmit experimentally and appeared to be only weakly contagious.

Journal of the Fisheries Research Board of Canada

Immunomodulation and disease resistance in postyearling rainbow trout infected with Myxobolus cerebralis, the causative agent of whirling disease

Myxobolus cerebralis , the myxosporean parasite that causes whirling disease, has a number of deleterious effects on its salmonid host. Although it is well established that juvenile salmonids in the active stages of whirling disease mount an immune response to the pathogen, the occurrence and longevity of any related immunomodulatory effects are unknown. In this study, postyearling rainbow trout Oncorhynchus mykiss infected with M. cerebralis were examined for leukocyte functions and for resistance to Yersinia ruckeri , a bacterial pathogen of salmonids. Compared with uninfected controls, M. cerebralis -infected fish showed lower proliferative lymphocyte responses to four mitogens (concanavalin A, pokeweed mitogen, phytohemagglutinin, and lipopolysaccharide). Conversely, M. cerebralis -infected fish displayed greater bactericidal activity of anterior kidney macrophages than did uninfected fish. After bath challenges with Y. ruckeri , M. cerebralis -infected fish had slightly lower survival and a more rapid onset of mortality than did the control fish. Renal tissue and fecal samples from M. cerebralis -infected and uninfected survivors were cultured for the presence of Y. ruckeri , and no difference in prevalence was noted between the two groups. Because immunomodulatory changes in the M. cerebralis -infected fish involved functional enhancement and suppression of different leukocyte populations, disease resistance among M. cerebralis -infected fish in the later stages of whirling disease will probably vary with the secondary pathogen and the nature of immune response the pathogen evokes.

Journal of Aquatic Animal Health

Pathophysiology of infectious hematopoietic necrosis virus disease in rainbow trout (Salmo gairdneri): early changes in blood and aspects of the immune Response after Injection of IHN Virus

Juvenile rainbow trout ( Salmo gairdneri ) were injected with infectious hematopoietic necrosis (IHN) virus and various hematological and blood chemical changes were monitored over 9 days. The packed cell volume, hemoglobin, red blood cell count, and plasma bicarbonate were significantly depressed by day 4. Plasma chloride, calcium, phosphorus, total protein, and blood cell types did not change during the 9 days. Furthermore, plasma LDH isozyme was significantly increased by the fourth day, and fish infected with infectious pancreatic necrosis virus, Vibrio anguillarum , Aeromonas salmonicida , and redmouth bacterium did not show specific LDH isozyme alterations. Acid-base alterations occurred at 10 C but not at 18 C. The acid-base imbalance and elevation of the LDH isozyme were consistently associated with the early development of the disease.The immune response after injection of IHN virus was determined and protection from disease was tested by passive immunization. Actively immunized fish developed IHN-neutralizing antibodies within 54 days after injection of virus, and the antibodies were protective when juvenile fish were passively immunized and experimentally challenged with IHN virus.

Journal of the Fisheries Research Board of Canada

Epidemiology of viral hemorrhagic septicemia (VHS) among juvenile Pacific herring and Pacific sandlances in Puget Sound, Washington

Viral hemorrhagic septicemia (VHS) and the associated virus (VHSV) were identified in newly metamorphosed Pacific herring Clupea pallasi and Pacific sand lances Ammodytes hexapterus captured from Puget Sound, Washington, between 1995 and 1998. During that 4-year period, virus was detected in less than 1% of free-ranging, age-0 Pacific herring; however, when groups of these fish were confined in the laboratory, they experienced severe mortality, occasionally exceeding 50%, with the prevalence of VHSV reaching 100% by 14 d postcapture. At 7–21 d postcapture, VHSV titers peaked in excess of 108 plaque-forming units/g of tissue; by 30 d postcapture, however, the virus could no longer be isolated. Fish surviving beyond 30 d eliminated the virus from their tissues, but some remained lethargic and continued to show signs of hemorrhage around the mouth, skin, and fins until about 6 weeks postcapture. No cutaneous ulcers were observed during either the acute or the recovery phases of infection. Eighteen-month-old Pacific herring captured from the same area were also negative for VHSV but developed active infections after confinement for 7 d. Unlike younger fish, only 8.4% of these older fish died of VHS, and 7.7% of survivors were positive for VHSV at 7–10 d postcapture, which suggests that a higher proportion of the older fish had developed resistance to VHSV from prior exposure to it. Three months after fatalities ceased in the laboratory-held fish, the surviving fish were challenged with 5 3 103 plaque-forming units/mL of VHSV for 1 h. No mortality was observed during the next 30 d, and virus was recoverable at very low titers in fewer than 5% of the challenged fish, indicating the development of an active immunity to VHSV. Laboratory cohabitation of infected wild Pacific herring with laboratory-reared, specific-pathogen-free Pacific herring resulted in transmission of VHSV to the nonimmune fish, with the resulting course of disease resembling that seen in wild Pacific herring confined in the laboratory. The possible effects of VHS on stocks of Pacific herring are discussed.

Journal of Aquatic Animal Health

Resistance and Protective Immunity in Redfish Lake Sockeye Salmon Exposed to M Type Infectious Hematopoietic Necrosis Virus (IHNV)

Differential virulence of infectious hematopoietic necrosis virus (IHNV) isolates from the U and M phylogenetic subgroups is clearly evident in the Redfish Lake (RFL) strain of sockeye salmon Oncorhynchus nerka. In these fish, experimental immersion challenges with U isolates cause extremely high mortality and M isolates cause low or no mortality. When survivors of M virus immersion challenges were exposed to a secondary challenge with virulent U type virus they experienced high mortality, indicating that the primary M challenge did not elicit protective immunity. Delivery of a moderate dose (2 × 104 plaque-forming units [PFU]/fish) of virus by intraperitoneal injection challenge did not overcome RFL sockeye salmon resistance to M type IHNV. Injection challenge with a high dose (5 × 106 PFU/fish) of M type virus caused 10% mortality, and in this case survivors did develop protective immunity against a secondary U type virus challenge. Thus, although it is possible for M type IHNV to elicit cross-protective immunity in this disease model, it does not develop after immersion challenge despite entry, transient replication of M virus to low levels, stimulation of innate immune genes, and development of neutralizing antibodies in some fish.

Journal of Aquatic Animal Health

Resistance of underyearling and yearling Atlantic salmon and lake trout to supersaturation with air

Juvenile Atlantic salmon Salmo salar and lake trout Salvelinus namaycush of different sizes were exposed for 96 h to water supersaturated with air. Underyearlings (Atlantic salmon shorter than 66 ± 8 mm [mean total length ± SD] and lake trout shorter than 96 mm) were resistant to high dissolved-gas pressures. The level of supersaturation causing 50% mortality was lower for yearling Atlantic salmon (148 ± 17 mm long) than for underyearling Atlantic salmon, and sensitivity to supersaturation was greater in lake trout 96 ± 1 to 153 ± 17 mm long than in those 23 ± 1 to 46 ± 4 mm long. Incidence of cutaneous emphysema and other signs of gas bubble disease increased as supersaturation increased. The number of bubbles and number of sites with bubbles were higher in the older fish. Among fish 150 ± 17 mm long, Atlantic salmon seemed to be less resistant to gas supersaturation than did lake trout.

Journal of Aquatic Animal Health

Oral immunization of rainbow trout ( Salmo gairdneri ) against an etiologic agent of "redmouth disease"

Rainbow trout were fed a pelleted diet containing killed cells of the etiologic agent of a bacterial disease, redmouth. These fish in addition to appropriate controls were subsequently challenged with virulent homologous organisms. Ninety per cent of the redmouth immunized fish survived the basic challenge using virulent organisms in contrast to 20% survival for the controls. Multiple challenge doses at increased levels also are discussed.

Journal of the Fisheries Research Board of Canada

Immunization of fishes: A review

Immune response in fishes, which are ectothermic animals, depends on temperature. The optimum response is at the optimum temperature for the species. It is, however, slower in coldwater fishes, such as salmonids, and faster in warmwater fishes. Serum of fishes contains proteins very similar, but not identical, with those of mammals. Immune bodies are contained in gamma, beta, and alpha globulins. Gamma globulin is absent in some fishes. Fishes can be effectively immunized by injection of antigens, however, this is not a practical method. For this reason oral immunization has been attempted repeatedly but the effectiveness is variable.

Journal of Wildlife Diseases

Swimming endurance of bull trout, lake trout, arctic char, and rainbow trout following challenge with Renibacterium salmoninarum

We tested the swimming endurance of juvenile bull trout Salvelinus confluentus , lake trout S. namaycush , Arctic char S. alpinus , and rainbow trout Oncorhynchus mykiss at 9°C and 15°C to determine whether sublethal infection from a moderate challenge of Renibacterium salmoninarum administered months before testing affected the length of time fish could maintain a swimming speed of 5-6 body lengths per second in an experimental flume. Rainbow trout and Arctic char swam longer in trials than did bull trout or lake trout, regardless of challenge treatment. When we tested fish 14-23 weeks postchallenge, we found no measurable effect of R. salmoninarum on the swimming endurance of the study species except for bull trout, which showed a mixed response. We conducted additional trials with bull trout 5-8 weeks postchallenge to determine whether increasing the challenge dose would affect swimming endurance and hematocrit. In those tests, bull trout with clinical signs of disease and those exposed to the highest challenge doses had significantly reduced swimming endurance compared with unchallenged control fish. Fish hematocrit levels measured at the end of all swimming endurance tests varied among species and between test temperatures, and patterns were not always consistent between challenged and control fish.

Journal of Aquatic Animal Health

Effects of a low-lipid diet on the gut microbiome and head kidney transcriptome of juvenile Chinook Salmon

Objective Pacific salmon Oncorhynchus spp. reared in production hatcheries are typically fed high-lipid, energy-dense diets to achieve large size and high body condition prior to release. In contrast, juveniles in natural environments tend to consume low-lipid, high-protein diets, and fish reared for research or conservation purposes are sometimes fed diets that are formulated to mimic natural diets and promote wild-like phenotypes. Understanding how these alternative diets affect fish health beyond growth and body condition could ultimately contribute to improving hatchery fish fitness. Methods In this work, we evaluated changes in the fecal microbiome and gene expression of juvenile Chinook Salmon O. tshawytscha on a standard high-lipid hatchery diet versus a low-lipid diet formulated to mimic the nutrition profile of natural-origin fish. To evaluate the time scale at which diet alters the fecal microbiome, we collected longitudinal samples over a 12-week period and switched the diets of a subset of fish twice during the experiment. We used 16S ribosomal RNA gene amplicon sequencing to characterize fecal microbiome differences between fish on the two diets as well as hatchery-reared fish at a production hatchery, hatchery fish that had been captured after release into a stream, and natural-origin, stream-reared fish of similar ages. Additionally, we conducted RNA sequencing on head kidney samples from laboratory-reared fish to evaluate changes in gene expression in this important immune organ. Results We found that the low-lipid diet and the hatchery diet resulted in microbiomes that differed from the microbiome of natural-origin fish and from each other and that diet-driven changes to the microbiome could occur in under 14 d. The low-lipid diet did not result in a microbiome that resembled the microbiome of naturally produced fish. Instead, the low-lipid diet resulted in a microbiome community that was distinct from those of fish reared on the hatchery diet and fish sampled from the wild. The RNA sequencing results indicated differential enrichment of pathways related to immunity, metabolism, and hormone synthesis between fish that were fed the two experimental diets. Conclusions The results suggest that additional environmental factors influence the microbiome more strongly than diet formulation or that the low-lipid diet has a smaller effect on the microbiome than a natural, ­invertebrate-based diet. Given that the gut microbiome and systemic immune function contribute significantly to disease resistance, our findings highlight the importance of understanding how diets fed to fish in captivity may affect fish health beyond growth and body condition metrics.

Journal of Aquatic Animal Health

Nutritional factors in the biochemical pathology of Corynebacterial kidney disease in the coho salmon (Oncorhynchus kisutch)

The influence of diet ingredient on the morbidity and biochemical pathogenesis of corynebacterial kidney disease was investigated using juvenile coho salmon ( Oncorhynchus kisutch ) fed the Abernathy dry ration made up with either corn gluten or cottonseed meal (isoprotein, isocaloric substitution). Evaluation of incidence of infection, pituitary activation and aspects of carbohydrate metabolism, acid-base balance, renal function, and hematopoietic activity showed that the actual disease incidence was about the same for both diets but the nonspecific stress of infection was more severe in fish fed the corn gluten.Discriminant function calculations combining four physiological parameters gave a probability of 0.86 for successfully diagnosing infected fish on the basis of these blood chemistry tests.

Journal of the Fisheries Research Board of Canada

Comparative sensitivities of diagnostic procedures used to detect bacterial kidney disease in salmonid fishes

Kidney and spleen homogenates from each of 60 coho salmon ( Oncorhynchus kisutch ) and steelhead trout ( Salmo gairdneri ) were examined for detection of Renibacterium salmoninarum . The proportions of positives differed widely with the detection procedures used: in coho salmon, 5% were positive by the Gram-stain procedure, 10% by the direct fluorescent antibody test, 48% by bacteriological isolation, 65% by staphylococcal coagglutination, and 73% by counterimmunoelectrophoresis; in steelhead trout, 3% were positive by Gram-stain, 8.3% by fluorescent antibody, 17% by bacteriological isolation, and 67% by counterimmunoelectrophoresis. Renibacterium salmoninarum was not detected in either coho salmon or steelhead trout by immunodiffusion analysis.

Journal of Wildlife Diseases

Quarantine of Aeromonas salmonicida-harboring ebonyshell mussels (Fusconaia ebena) prevents transmission of the pathogen to brook trout (Salvelinus fontinalis)

Furunculosis, caused by the bacterium Aeromonas salmonicida , was artificially induced in brook trout ( Salvelinus fontinalis ) in an experimental tank. Ebonyshells ( Fusconaia ebena ) were placed to cohabit with these fish to acquire the pathogen through siphoning. After 2 wk of cohabitation, 10 of the mussels were assayed by bacterial culture and all were found to harbor A. salmonicida . The mean cell count from soft tissue homogenates was 1.84 × 10 5 cfu/g, which comprised an average 14.41% of the total bacteria isolated from tissues. From the fluids, a mean of 2.84 × 10 5 A. salmonicida cfu/mL was isolated, which comprised an average of 17.29% of the total bacterial flora. The mussels were removed from the cohabitation tank and distributed equally among five previously disinfected tanks, 35 per tank. The F. ebena in each tank were allowed to depurate A. salmonicida for various durations: 1, 5, 10, 15 or 30 days. After each group had depurated for their assigned time, 10 were assayed for bacteria, tank water was tested, and 20 pathogen-free bioindicator brook trout were added to cohabit with the remaining mussels. Depuration was considered successful if A. salmonicida was not isolated from tank water or the mussels, and there was no infection or mortality to bioindicator fish. After 1 day of depuration, A. salmonicida was not isolated from the soft tissues; however, it was isolated from one of the paired fluids (10% prevalence). The tank water tested positive, and the bioindicator fish became infected and died. From the 5-day depuration group, A. salmonicida was not isolated from soft tissues, but was isolated from three fluids (30%; mean = 1.56 × 10 2 cfu/mL). Tank water from the 5-day group was negative, and there was no mortality among the bioindicator fish. However, A. salmonicida was isolated from 2 of 20 fish at the end of the 14-day observation period. One F. ebena fluid sample was positive for A. salmonicida from the 10-day depuration group, but none of the soft tissue homogenates. The pathogen was not isolated from 10-day tank water, but there was a 30% cumulative mortality to the bioindicator fish. Aeromonas salmonicida was not isolated from any of the soft tissue homogenates, fluids or tank water from the 15 day or 30 day depuration groups, and the bioindicator fish remained pathogen- and disease-free. Study results showed that the F. ebena were harboring a high A. salmonicida cell load going into depuration, but at 15 days and beyond, the pathogen had been depurated to the extent that the mussels did not serve as pathogen vectors.

Journal of Shellfish Research

Isolation and phenotypic characterization of an oxidase-negative Aeromonas salmonicida causing furunculosis in coho salmon (Oncorhynchus kisutch)

An oxidase-negative Aeromonas salmonicida was isolated from coho salmon ( Oncorhynchus kisutch ) suffering from an epizootic of furunculosis at the state hatchery near Belfair, Washington. Typical, oxidase-positive A. salmonicida was isolated concurrently from the same population of fish. Mortality was controlled with medicated feed treatments. Evidence supporting the identification of the two types of A. salmonicida is presented. Methods for the proper identification of oxidase-negative A. salmonicida isolates are evaluated.

Journal of Wildlife Diseases

Immunization of salmonids against Yersinia ruckeri: Significance of humoral immunity and cross protection between serotypes

Brook trout ( Salvelinus fontinalis ) were immunized with bacterins containing either Serotype 1 or 2 isolates of Yersinia ruckeri to determine the relative degree of cross-protection afforded when the fish were challenged with the homologous or heterologous serotype. While fish immunized with pH-lysed bacterins produced highly specific agglutinins that did not cross-react with antigens derived from a heterologous serotype of Y. ruckeri all fish were protected against experimental challenge, regardless of which serotype was used for bacterin production and experimental challenge. Other experiments indicated that brook trout injected intraperitoneally with highly specific antibodies could not be passively immunized against experimental challenge.

Journal of Wildlife Diseases