Effects of downstream migration on myosin heavy chain expression and dihydropyridine receptor density in farmed smolt of Atlantic salmon

The expression of myosin heavy chain (MHC) subunits and dihydropyridine receptors (DHPRs) from red and white tail muscles of cultured smolts of Atlantic salmon Salmo salar was analysed from samples taken: (1) before the fish were transferred to the river and (2) after the migration distance of c . 50 km. The relative work load of migrating fish, estimated on the basis of swimming speed of Atlantic salmon and discharge rate of the River Simojoki, Finland, was maximally 1700 times higher than that of fish in culture. The data show that after five migration days the density of DHPRs in the muscles of the smolts was increased already. Furthermore, the results indicated a transition from the fast‐to‐slow MHC isoform. Transfer of fish to a natural environment and downstream migration thus had a significant effect on the expression of muscle proteins.     

Distribution,prevalence and severity of Parvicapsula minibicornis infections among anadromous salmonids in the Fraser River,British Columbia,Canada

Polymerase chain reaction (PCR) and microscopic examination of stained kidney sections were used to diagnose infections with the myxozoan parasite Parvicapsula minibicornis in maturing Fraser River salmon. In 2 series of collections, the parasite was detected in 109 of 406 migrating sockeye salmon Oncorhynchus nerka belonging to Early Stuart, Early Summer and Summer run-timing groups, mainly upper Fraser River stocks. However, the parasite was detected neither in fish at sea nor once they had migrated several 100 km upstream. Prevalence then increased to 95% or greater at the spawning grounds. Histological examination of kidney was less sensitive than PCR in detecting the parasite in salmon collected from the earliest sites in both collections found positive by PCR. Severity of infection was greatest at the spawning grounds. Development of infection in sockeye, measured by prevalence, severity or by the rate of false-negative histological diagnoses, appeared to be a useful estimate of in-river residence time. Prevalence and severity of infections in sequential samples of Harrison River and Weaver Creek sockeye stocks collected from the Harrison River indicated that more time had elapsed since parasite transmission than would be predicted based on migration distance alone. Pink salmon Oncorhynchus gorbuscha, coho salmon O. kisutch and chinook salmon O. tshawytscha were found to be infected with the parasite. Development of P. minibicornis in pink salmon was most similar to that in sockeye. Pink and coho salmon may be at risk to the pathological consequences of P. minibicornis infection.     

Changes in plasma protein patterns in smolting Atlantic salmon, Salmo salar L., are not dependent on changed growth rates

Juvenile Atlantic salmon, Salmo salar L., were raised under simulated natural photoperiod (SNP) and constant light (CL). SNP fish displayed typical characteristics of smolts in the spring including changes in colouration, elevated gill Na⁺/K⁺ ATPase activity and ability to pass a seawater challenge. CL fish grew at rates comparable to SNP fish, but did not demonstrate any of these molt characteristics.
Electrophoretic analyses of plasma proteins from both groups of fish revealed that SNP fish displayed reproducible qualitative changes in protein patterns from February to May with May patterns being the most complex. CL fish displayed patterns similar to SNP fish. The timing of the appearance of patterns was influenced by photoperiod as indicated by the fact that CL fish had a progression of pattern changes retarded by at least one month relative to SNP fish. The changes in plasma protein patterns in smolting Atlantic salmon were not dependent on changed growth rates.     

Elevation of plasma cortisol during the spawning migration of landlocked kokanee salmon (Oncorhynchus nerka kennerlyi)

Kokanee salmon (Oncorhynchus nerka kennerlyi ), a landlocked subspecies of sockeye salmon, exhibited hypothalamic-pituitary interrenal (HPI, adrenal homologue) axis activation and an increase in plasma cortisol concentration up to 639 +/- 55.9 ng/ml in association with upstream migration in the upper Colorado River even though they were not exposed to a change in salinity and lengthy migration. Kokanee salmon were collected at various stages of migration and concomitant sexual maturation. The pattern of cortisol elevation in kokanee is similar to that in ocean-run sockeye salmon (O. nerka nerka). The presence of plasma cortisol elevation in an upstream migrating, landlocked Pacific salmon suggests that stressors previously considered to cause the cortisol increase, such as long-distance migration and changes in salinity, may not be primary causes of the HPI axis activation.     

Factors influencing the survival of outmigrating juvenile salmonids through multiple dam passages: an individual‐based approach

Substantial declines of Pacific salmon populations have occurred over the past several decades related to large‐scale anthropogenic and climatic changes in freshwater and marine environments. In the Columbia River Basin, migrating juvenile salmonids may pass as many as eight large‐scale hydropower projects before reaching the ocean; however, the cumulative effects of multiple dam passages are largely unknown. Using acoustic transmitters and an extensive system of hydrophone arrays in the Lower Columbia River, we calculated the survival of yearling Chinook salmon (Oncorhynchus tshawytscha) and steelhead (O. mykiss) passing one, two, or three dams. We applied a unique index of biological characteristics and environmental exposures, experienced by each fish individually as it migrated downstream, in order to examine which factors most influence salmonid survival. High outflow volumes led to involuntary spill in 2011 and created an environment of supersaturated dissolved gas concentrations. In this environment, migrating smolt survival was strongly influenced by barometric pressure, fish velocity, and water temperature. The effect of these variables on survival was compounded by multiple dam passages compared to fish passing a single dam. Despite spatial isolation between dams in the Lower Columbia River hydrosystem, migrating smolt appear to experience cumulative effects akin to a press disturbance. In general, Chinook salmon and steelhead respond similarly in terms of survival rates and responses to altered environmental conditions. Management actions that limit dissolved gas concentrations in years of high flow will benefit migrating salmonids at this life stage.     

Commercial blast-freezing of third-stage Anisakis simplex larvae encapsulated in salmon and rockfish

Sixty-four fish were blast-frozen to -35 C for 15 hr to determine the effects of commercial blast-freezing on the viability of third-stage larvae of Anisakis simplex encapsulated in the muscle and viscera of sockeye salmon (Oncorhynchus nerka) and canary rockfish (Sebastes pinniger). Parallel tests were conducted on larval nematodes in 16 whole (round) salmon, 16 dressed salmon (heads and viscera removed), and 32 whole (round) rockfish. After blast-freezing, 4 in-the-round salmon, 4 dressed salmon, and 8 in-the-round rockfish were examined at 1, 24, 48, and 72 hr. A total of 3,539 dead and 6 live larvae were collected from the fish tissues after standard enzymatic digestion. Salmon were infected with 1,245 of these larvae, and rockfish with 2,300. The 6 live worms, 2 from salmon and 4 from rockfish rounds, were recovered from muscle 1 hr after freezing; they were slightly motile and showed severe internal damage. No viable worms were found at or after 24 hr. The commercial blast-freezing process effectively killed larval nematodes in whole or dressed fish. Market-ready samples of previously blast-frozen silver salmon (O. kisutch) and chum salmon (O. keta) fillets and chum salmon steaks yielded no live worms, thereby confirming the efficacy of this process.     

Population Dynamics of the Pink Salmon Oncorhynchus gorbuschain the Basin of the Sea of Okhotsk in the 1990s

In the 1990s, an extensive body of data was gathered on the size of the Oncorhynchus gorbuschapink salmon populations of the Sea of Okhotsk at all the main developmental stages. A significant increase in numbers was found for juvenile pink salmon migrating into the offshore regions of the Sea of Okhotsk and the Pacific waters around the Kurils: from 250–450 million in 1990–1991 to 807–1570 million fish in 1993–1999. The overall number of migrating pink salmon in even years sharply increased in 1994 up to 215 million fish. After 1994, this estimate exceeded the number of migrating pink salmon in preceding odd years. Ocean survival of juvenile pink salmon gradually declined. This may be related to changes in the North Pacific pelagic ecosystems.     

Changes in Atlantic salmon (Salmo salar) epidermal mucus protein composition profiles following infection with sea lice (Lepeophtheirus salmonis)

The mucus protein profile of Atlantic salmon (Salmo salar) and changes due to infection with sea lice (Lepeophtheirus salmonis) were examined. Two-dimensional gel electrophoresis was performed on salmon skin mucus and comparisons between control and infected fish mucus were made. LC MS/MS identified intracellular proteins, calmodulin, actin, and hemopexin and plasma proteins, such as apolipoproteins, lectin, plasminogen and transferrin. Plasma proteins in the mucus may result from either direct expression by epidermal cells, leakage of plasma or via a secondary circulation system. Therefore, RT-PCR was used to measure mRNA of transferrin and lectin in Atlantic salmon skin. Transferrin expression was observed suggesting direct expression by the epidermis. Lectin expression was not detected suggesting another mechanism of entry into mucus, either leakage from plasma or secondary circulation. The lack of observable albumin on 2D gels, suggests that mucus lectin may arise from the secondary circulation route. Interestingly, β-actin was a significant component of Atlantic salmon mucus. Cleaved actin and transferrin fragments were observed and positively correlated with sea lice infection suggestive of proteolytic activity. Increased levels of cleaved transferrin during sea lice infection may activate the nitrous oxide response of salmon macrophages, as part of the fish's immune response to sea lice infection.     

Enzymes released from Lepeophtheirus salmonis in response to mucus from different salmonids

Adult and mobile preadult sea lice Lepophtheirus salmonis were incubated with mucus samples from rainbow trout (Oncorhynchus mykiss), coho salmon (O. kisutch), Atlantic salmon (Salmo salar), and winter flounder (Pseudopleuronectes americanus) to determine the response of L. salmonis to fish skin mucus as assessed by the release of proteases and alkaline phosphatase. There was variation in the release of respective enzymes by sea lice in response to different fish. As well, sealice collected from British Columbia responded differently than New Brunswick sea lice to coho salmon mucus. Fish mucus and seawater samples were also analyzed using protease gel zymography to observe changes in the presence of low molecular weight (LMW) proteases after L. salmonis incubation. Significantly higher proportions of sea lice secreted multiple bands of L. salmonis-derived LMW proteases after incubation with rainbow trout or Atlantic salmon mucus in comparison with seawater, coho salmon, or winter flounder mucus. Susceptibility to L. salmonis infections may be related to the stimulation of LMW proteases from L. salmonis by fish mucus. The resistance of coho salmon to L. salmonis infection may be due to agents in their mucus that block the secretion of these LMW proteases or factors may exist in the mucus of susceptible species that stimulate their release.     

The use of chemical cues in the upstream migration of coho salmon, Oncorhynchus kisutch Walbaum

The migratory behaviour of coho salmon was studied in the field. Coho salmon, Oncorhynchus kisutch , exposed to synthetic odours in a hatchery and released into Lake Michigan as juveniles, were recaptured as migrating adults 18 months later. The movements of 43 individuals were followed in experiments examining the role of imprinted chemical cues in the homing migration. Movements in relation to controlled distributions of the imprinting odours revealed that chemical cues are used by the salmon in their upstream migration. Acting with the rheotactic response, fish were guided upstream by imprinting odours simulating their home stream. Coho salmon, in the absence of their imprinting odour, respond negatively to the current. Differential responses to current appear to serve in segregating fish imprinted to odours of different home streams.     

Changes in hydrolytic enzyme activities of naïve Atlantic salmon Salmo salar skin mucus due to infection with the salmon louse Lepeophtheirus salmonis and cortisol implantation

The changes in the activities of mucus hydrolytic enzymes and plasma cortisol levels were examined following infection of Atlantic salmon Salmo salar with the salmon louse Lepeophtheirus salmonis and these changes were compared with those resulting from elevated plasma cortisol. Salmon were infected at high (Trial 1; 178 +/- 67) and low (Trial 2; 20 +/- 13) numbers of lice per fish and the activities of proteases, alkaline phosphatase, esterase and lysozyme in the mucus, as well as plasma cortisol levels were determined. At both levels of infection, there were significant increases of protease activity over time (1-way K-WANOVA; Trial 1, p = 0.004; Trial 2, p < 0.001). On several sampling days, generally on later days in the infections, the mucus protease activities of infected fish were significantly higher than control fish (Student's t-tests; p < 0.05). In addition, zymography experiments demonstrated bands of proteases at 17 to 22 kDa in the mucus of infected salmon that were absent in the mucus from non-infected fish and absent in the plasma of salmon. The intensity of these protease bands increased in the mucus over the course of both infections. However, plasma cortisol levels were elevated only in the heavily infected fish from the first trial. At high infection levels (Trial 1), alkaline phosphatase activity was higher in the mucus of infected fish at all days (t-test, p < 0.05). However, at the lower infection level (Trial 2), the mucus alkaline phosphatase activity did not differ significantly between infected and non-infected fish. Esterase and lysozyme activities were very low and did not change with time nor between non-infected and infected salmon in either challenge. Mucus enzyme activities of cortisol-implanted salmon did not change over time, nor were there any differences in activities between cortisol-implanted and control salmon. The present study demonstrates biochemical changes resulting from sea lice infection of Atlantic salmon occurring at the site of host-pathogen interaction, the mucus layer. However, the origin of these enzymes, whether host or pathogen, remains to be determined.     

Physiological and molecular endocrine changes in maturing wild sockeye salmon, <Emphasis Type="Italic">Oncorhynchus nerka</Emphasis>, during ocean and river migration

Maturing adult sockeye salmon Oncorhynchus nerka were intercepted while migrating in the ocean and upstream in freshwater over a combined distance of more than 1,300 km to determine physiological and endocrine changes associated with ionoregulation. Sockeye migrating through seawater and freshwater showed consistent declines in gill Na⁺/K⁺-ATPase (NKA) activity, plasma osmolality and plasma chloride concentration. In contrast, plasma sodium concentration became elevated in seawater as fish approached the river mouth and was then restored after sockeye entered the river. Accompanying the movement from seawater to freshwater was a significant increase in mRNA for the NKA α1a subunit in the gill, with little change in the α1b subunit. Potential endocrine signals stimulating the physiological changes during migration were assessed by measuring plasma cortisol and prolactin (Prl) concentrations and quantifying mRNA extracted from the gill for glucocorticoid receptors 1 and 2 (GR1 and GR2), mineralocorticoid receptor (MR), growth hormone 1 receptor (GH1R), and prolactin receptor (PrlR). Plasma cortisol and prolactin concentrations were high in seawater suggesting a preparatory endocrine signal before freshwater entry. Generally, the mRNA expression for GR1, GR2 and MR declined during migration, most notably after fish entered freshwater. In contrast, PrlR mRNA increased throughout migration, particularly as sockeye approached the spawning grounds. A highly significant association existed between gill PrlR mRNA and gill NKA α1a mRNA. GH1R mRNA also increased significantly, but only after sockeye had migrated beyond tidal influence in the river and then again just before the fish reached the spawning grounds. These findings suggest that cortisol and prolactin stimulate ionoregulation in the gill as sockeye salmon adapt to freshwater.     

Resurrecting an extinct salmon evolutionarily significant unit: archived scales, historical DNA and implications for restoration

Archival scales from 603 sockeye salmon (Oncorhynchus nerka), sampled from May to July 1924 in the lower Columbia River, were analysed for genetic variability at 12 microsatellite loci and compared to 17 present‐day O. nerka populations—exhibiting either anadromous (sockeye salmon) or nonanadromous (kokanee) life histories—from throughout the Columbia River Basin, including areas upstream of impassable dams built subsequent to 1924. Statistical analyses identified four major genetic assemblages of sockeye salmon in the 1924 samples. Two of these putative historical groupings were found to be genetically similar to extant evolutionarily significant units (ESUs) in the Okanogan and Wenatchee Rivers (pairwise F_ST = 0.004 and 0.002, respectively), and assignment tests were able to allocate 77% of the fish in these two historical groupings to the contemporary Okanogan River and Lake Wenatchee ESUs. A third historical genetic grouping was most closely aligned with contemporary sockeye salmon in Redfish Lake, Idaho, although the association was less robust (pairwise F_ST = 0.060). However, a fourth genetic grouping did not appear to be related to any contemporary sockeye salmon or kokanee population, assigned poorly to the O. nerka baseline, and had distinctive early return migration timing, suggesting that this group represents a historical ESU originating in headwater lakes in British Columbia that was probably extirpated sometime after 1924. The lack of a contemporary O. nerka population possessing the genetic legacy of this extinct ESU indicates that efforts to reestablish early‐migrating sockeye salmon to the headwater lakes region of the Columbia River will be difficult.     

Evaluation of biochemical methods for the non‐destructive identification of sex in upstream migrating salmon and sea trout

Female‐specific markers of reproductive activity [plasma 17β‐oestradiol (E2), vitellogenin (VTG) and alkali‐labile phosphoprotein phosphorous (ALP)] were measured over 12 months in a captive population of brown trout Salmo trutta . During the early months of the reproductive season (February to May) and using the concentration of plasma E2 or plasma ALP as a marker for females the proportion of fish in which sex was misidentified was high (15–50%). The misidentification rate was considerably lower (1–8%) using plasma VTG. Preliminary evaluation of a commercial immunochromatographic VTG test system as a screen for the presence or absence of VTG in plasma from brown trout provided results that were consistent with those obtained from direct measurement of plasma VTG levels by enzyme‐linked immunosorbent assay (ELISA). These preliminary conclusions were verified by sampling upstream‐migrating anadromous brown trout, sea trout, and Atlantic salmon Salmo salar trapped over a 6 month period. Plasma E2 levels did not satisfactorily discriminate between male and female sea trout and Atlantic salmon. Plasma VTG levels in both species, however, were bimodally distributed and it was assumed that this divergence corresponded to male (plasma VTG levels <10 μg ml⁻¹) and female (plasma VTG levels >800 μg ml⁻¹) fishes. Plasma ALP provided a more accurate indication of sex in the wild Atlantic salmon and sea trout than was suggested by the pilot study on captive brown trout. The commercial immunochromatographic VTG test system provided results that were wholly consistent with the data obtained from the trapped fishes by direct measurement of plasma VTG.     

Physiological seawater adaptation in juvenile Atlantic salmon (Salmo salar) autumn migrants

Summary

• 1 About 25 % of juvenile Atlantic salmon (Salmo salar) migrating downstream in the River Frome in southern England do so in the autumn rather than in the spring. Here, we examine the physiological status of these fish with regard to those features that adapt them to sea water during the parr–smolt transformation (i.e. gill Na⁺K⁺ ATPase activity; the number, size and type of chloride cells on the gill lamellae; salinity tolerance and relative plasma thyroid levels).
• 2 Autumn migrants, and those fish which subsequently reside in the tidal reaches during the winter, are not sufficiently physiologically adapted to permit permanent or early, entry into the marine environment.
• 3 It is not known what proportion of autumn migrating fish survive and return to spawn as adults. If significant numbers do return, however, the production from tidal reach habitats must be taken into account in the development of salmon stock management strategies, especially monitoring and assessment programmes, and in the evaluation of factors affecting stocks.

     

Development and application of a nested PCR to monitor brood stock salmonid ovarian fluid and spleen for detection of the fish pathogen Flavobacterium psychrophilum

AIMS: To develop a nested PCR to detect Flavobacterium psychrophilum based on the intergenic spacer region 16S-23S rRNA and in 16S rRNA for analysis of brood stock salmonid fish samples. METHODS AND RESULTS: The sensitivity and specificity of the test was evaluated using pure cultures, spiked and naturally contaminated samples. Samples were internal organs (spleen and kidney), eggs and ovarian fluid from rainbow trout and coho salmon from European fish farms (France, Spain). This nested PCR was more specific and sensitive that the nested PCR based on 16S rRNA sequences primers only. The detection limit of this PCR assay was one bacterium per PCR tube corresponding to 10 bacteria/mg of spleen and 5 bacteria/ml from ovarian fluid. Analysis of mixed ovarian fluid samples from reproductive salmonids in various French hatcheries demonstrated that 69% of hatcheries were contaminated with Fl. psychrophilum. The analysis of individual samples demonstrated that 39% of rainbow trout (Oncorhynchus mykiss) and 62.5% of coho salmon (O. kisutch) samples were contaminated. CONCLUSIONS: The results demonstrated a very sensitive and specific detection of this fish pathogen and that most of the female rainbow trout and coho salmon breeders analysed carry Fl. psychrophilum in the ovarian fluid. SIGNIFICANCE AND IMPACT OF THE STUDY: The understanding of Fl. psychrophilum dissemination and transmission and the detection of asymptomatic carriers is important for the development of free breeders stock and for significantly decreasing Flavobacteriose.     

Changes in serum thyroxine levels during smolting in captive and wild Atlantic salmon, Salmo salar L.

On a commercial fish farm in Western Scotland, smolting Atlantic salmon, Salmo salar L., showed no major changes in serum thyroxine (T4) levels in the spring. In contrast, serum T4 levels in smolting, wild salmon, trapped as they left the Girnock Burn, Aberdeenshire, varied markedly over this period. Serum T4 levels were correlated with the stream discharge rate pertaining as the fish left the stream. Levels in migrating, smolting fish were higher than those in fish captured above the fish-trap by electro-fishing. In a stream channel, serum T4 levels in smolting fish caught previously in the fish-trap were rapidly elevated by exposure to increased water-flow, whereas levels in fish previously captured above the trap remained substantially unchanged. Serum T4 levels in salmon parr leaving the stream in autumn were generally lower than those in smolting fish leaving in spring and were not demonstrably affected by stream discharge rate.     

Development and characterization of a competitive polyclonal antibody enzyme-immunoassay for salmon insulin-like growth factor-II

This paper describes the development and validation of a competitive, polyclonal antibody enzyme-immunoassay (EIA) for the measurement of salmon and trout insulin-like growth factor-II (IGF-II). A polyclonal antiserum was raised against a synthetic peptide epitope, corresponding to amino acid residues 1-9 of the N-terminus of mature Atlantic salmon (Salmo salar) IGF-II. The antiserum was purified by hydrophobic charge induction chromatography (HCIC). The partially purified immunoglobulins were used in an enzyme-immunoassay system (EIA) resulting in a highly specific assay for salmon IGF-II with cross-reactivity of less than 0.01% for recombinant salmon IGF-I and recombinant salmon growth hormone (GH), and 5.57% for salmon insulin (sIns). The recombinant salmon IGF-II (rsIGF-II) standard curve limit of detection was 1.37 ng/ml with an EC(50) of 44.97+/-0.82 ng/ml. Intra- and interassay coefficients of variation were determined at 7.47% (n=15) and 7.42% (n=15), respectively. Added rsIGF-II was adequately recovered from acid-treated Atlantic salmon and rainbow trout (Oncorhynchus mykiss) plasma samples. Parallel dose-response inhibition curves were demonstrated for the plasma of both fish species tested. Circulating IGF-II levels of 22.26+/-2.66 and 18.24+/-1.43 ng/ml were determined for acid-treated plasma of normal adult Atlantic salmon and rainbow trout, respectively. This EIA should prove to be useful in the study of factors which influence circulating plasma levels of IGF-II in these fish species.