Identification of the Y chromosome in chinook salmon (Oncorhynchus tshawytscha)

The Y chromosome in chinook salmon, Oncorhynchus tshawytscha, was identified using fluorescence in situ hybridization (FISH) with a probe to a male-specific repetitive sequence isolated from this species. The probe highlights the distal end of the short arm of an acrocentric chromosome with a DAPI-bright interstitial band of variable size. The proximal portion of the short arm of the Y chromosome contains 5S rDNA sequences, which are also found on the short arms of six other acrocentric chromosomes in this species.     

A genetic linkage map for coho salmon (Oncorhynchus kisutch)

Construction of genetic linkage maps is an important first step for a variety of genomic applications, such as selective breeding in aquaculture, comparative studies of chromosomal evolution and identification of loci that have played key roles in the evolution of a species. Here we present a sex-specific linkage map for coho salmon. The map was constructed using 148 AFLP markers, 133 microsatellite loci and the phenotypic locus SEX . Twenty-four linkage groups spanning 287.4 cM were mapped in males, and 33 linkage groups spanning 429.7 cM were mapped in females. Several male linkage groups corresponded to two female linkage groups. The combination of linkage groups across both sexes appeared to characterize regions of 26 chromosomes. Two homeologous chromosomes were identified based on information from duplicated loci. Homologies between the coho and rainbow trout maps were examined. Eighty-six loci were found to form common linkage relationships between the two maps; these relationships provided evidence for whole-arm fissions, fusions and conservation of chromosomal regions in the evolution of these two species.     

Characterization of coho salmon (Oncorhynchus kisutch) insulin

Insulin has been isolated from islet tissue of coho salmon (Oncorhynchus kisutch) by gel filtration and HPLC and the complete amino acid sequence has been determined. The sequence differs from bovine insulin at 14 sites but all interchanges are conservative from the viewpoint of preservation of conformation. A comparison of insulin sequences from other fish is presented. Salmon insulin cross-reacts very weakly with antiserum to bovine insulin and vice versa. A completely homologous radioimmunoassay has been developed and used to estimate the insulin in salmon islet tissue and in plasma. The hypoglycemic effect of salmon insulin in salmon was more pronounced and persisted longer than that caused by identical doses of bovine insulin.     

Identification of the sex chromosome pair in chum salmon (Oncorhynchus keta) and pink salmon (Oncorhynchus gorbuscha)

Fluorescence in situ hybridization (FISH) using a probe to the male-specific GH-Y (growth hormone pseudogene) was used to identify the Y chromosome in the karyotypes of chum salmon (Oncorhynchus keta) and pink salmon (Oncorhynchus gorbuscha). The sex chromosome pair is a small acrocentric chromosome pair in chum salmon and the smallest metacentric chromosome pair in pink salmon. Both of these chromosome pairs are morphologically different from the sex chromosome pairs in chinook salmon (Oncorhynchus tshawytscha) and coho salmon (Oncorhynchus kisutch). The 5S rRNA genes are on multiple chromosome pairs including the sex chromosome pair in chum salmon, but at the centromeres of two autosomal metacentric pairs in pink salmon. The sex chromosome pairs and the chromosomal locations of the 5S rDNA appear to be different in all five of the North American Pacific salmon species and rainbow trout. The implications of these results for evolution of sex chromosomes in salmonids are discussed.     

The blood vasculature of the gastrointestinal tract in chinook, Oncorhynchus tshawytscha (Walbaum), and coho, O. kisutch (Walbaum), salmon

The venous and arterial vasculature of the chinook and coho salmon gastrointestinal tract were examined using corrosion casts and India ink injection techniques. Observations derived from 28 individuals of various sizes and of both sexes were used to construct simplified venous and arterial plans. Examination of the blood vasculature revealed the presence of a variety of anastomoses hitherto undescribed in teleosts.     

Ontogeny of the olfacto-retinalis projection in coho salmon (Oncorhynchus kisutch)

Filling of nucleus olfacto-retinalis neurons by cobaltous lysine injections into one eye of coho salmon of different ages revealed that the terminal nerve projection to the retina is established when the fish leave their freshwater environment. At this time salmons go through a metamorphosis termed "smoltification." FMRF-amide-like immunoreactive neurons in the nucleus olfacto-retinalis already exist in pre-smolts before the retino-petal projection is established. Thus, for the first time in any vertebrate, a projection of the terminal nerve is shown to develop during an advanced ontogenetic state.     

Glacial biogeography of North American coho salmon (Oncorhynchus kisutch)

To study the glacial biogeography of coho we examined 20 microsatellite loci and mitochondrial DNA D-loop sequence in samples from Alaska to California. Microsatellite data divided our samples among five biogeographic regions: (1) Alaska and northern coastal British Columbia; (2) the Queen Charlotte Islands; (3) the mainland coast of British Columbia and northern Washington State; (4) the Thompson River; and (5) Oregon and California. The D-loop sequence data suggested three geographical regions: (1) Oregon and California; (2) the Thompson River; and (3) all the other sites north of the southern ice margin. Microsatellite data revealed no difference in the number of alleles in different regions, but mitochondrial DNA data revealed a cline of decreasing diversity from south to north. We suggest that the two signals presented by these different marker types illuminate two time frames in the history of this species. Endemic microsatellite diversity in Alaska and on the Queen Charlotte Islands provides evidence in favour of Fraser Glaciation refugia in these regions. The loss of mitochondrial variation from south to north suggests that one of the earlier, more extensive, Pleistocene glaciations eliminated coho from its northern range.     

A single-locus minisatellite discriminates chinook salmon (Oncorhynchus tshawytscha) populations

A knowledge of genetic structure in natural populations is often necessary for conservation and management purposes, especially in declining Pacific salmon populations. To test for genetic differentiation between nine populations of chinook salmon, Oncorhynchus tshawytscha, from south-western British Columbia, Canada, DNA was extracted from 603 fish and hybridized with a single-locus minisatellite probe. Multivariate statistical analyses of the resulting allele size data permitted successful overall population identification of 52% (individual population range: 24–78%; P < 0.005), indicating a high level of genetic differentiation among the nine populations. Two of the nine populations were further analysed using data from a second minisatellite locus. The discrimination success rate improved from 81.1% (one-locus analyses) to 90.0% (two-locus analyses), indicating the potential for greatly increased resolution gained by the addition of more loci. These results indicate that variation at minisatellite loci can be used for assessing population-level genetic structure, even with artificial gene flow.     

Plasma profiles of the sex steroids and gonadotropins in maturing female spring chinook salmon (Oncorhynchus tshawytscha)

Plasma testosterone concentrations were low through the spring and early summer, concentrations began rising in late July and reached maximum levels by ovulation in September. Plasma concentrations of 11-ketotestosterone were low throughout sexual maturation until ovulation when a significant increase occurred. Plasma androstendione and 17β-estradiol concentrations were high throughout sexual maturation, and decreased significantly at ovulation. Plasma 17α,20β-dihydroxy-4-pregnen-3-one concentrations were low throughout maturation, and increased significantly at ovulation. Plasma gonadotropin I concentrations paralleled those of estradiol and exceeded gonadotropin II levels prior to ovulation. Plasma concentrations of gonadotropin II were low throughout the spring and summer, increasing dramatically at ovulation.     

Ontogenetic variations in the type and size of prey consumed by juvenile coho,Oncorhynchus kisutch,and chinook,O. tshawytscha,salmon

Synopsis Stomach contents of juvenile coho,Oncorhynchus kisutch, and chinook,O. tshawytscha, salmon collected in purse seines off the coast of Washington and Oregon were examined for variations related to predator size. There was a general trend toward increasing consumption of fish with increasing body size, due mainly to the increase in northern anchovy biomass consumed by the larger salmon. Most of the major prey taxa showed significant differences among the size classes examined for both salmon species. There was a direct relationship between predator and prey size for both coho and chinook, but considerable variation was found in prey length consumed within each size class. Prey width did not provide as good a fit as prey length for either species. In general, coho consumed larger fish prey in relation to their body length than chinook but there were substantial differences by month or year of collection.     

Characterization of Phase I biotransformation enzymes in coho salmon (Oncorhynchus kisutch)

Wild stocks of Pacific salmon in the Northwestern United States have declined in recent years, and the major factors contributing to these losses include water pollution and loss of habitat. In salmon, sublethal chemical exposures may impact critical behaviors (such as homing, feeding, predator-avoidance) that are important for species survival. Therefore, understanding the potential for these species to biotransform organic compounds within sensitive target tissues such as liver, gills and olfactory region can help estimate or predict their susceptibility to pollutants. In this study, we used real-time quantitative polymerase chain reaction (Q-PCR), Western blotting, and catalytic assays to characterize the expression of Phase I biotransformation enzymes in coho salmon (Oncorhynchus kisutch), a sensitive species in the Pacific Northwest. Gene expression analysis using Q-PCR assays developed for coho genes revealed the presence of the predominant cytochrome P450 mRNAs (CYP1A, CYP2K1, CYP2M1, CYP3A27) in the olfactory rosettes and provided quantitative mRNA expression levels in coho liver and gills. Q-PCR analysis revealed relatively high expression of the major CYP isoforms in the liver and olfactory rosettes, which was generally confirmed by Western blotting. Extrahepatic CYP expression was generally higher in the olfactory rosettes as compared to the gills. Catalytic studies demonstrated functional CYP1A-dependent ethoxyresorufin-O-deethylase, CYP2-dependent pentoxyresorufin-O-dealkylase, CYP2K1-dependent testosterone 16beta-hydroxylase, and CYP3A27-dependent testosterone 6beta-hydroxylase activities in liver, but not at detectable levels in gills. In contrast, flavin-containing monooxygenase (FMO)-dependent thiourea S-oxidase activity was readily observed in the gills and was substantially higher than that observed in liver. Collectively, the results of this study suggest that the olfactory rosettes are important sites of extrahepatic biotransformation in coho salmon, and that tissue specific-differences in Phase I metabolism may lead to contrasting tissue-specific biotransformation capabilities in this species.     

Outbreaks of phaeohyphomycosis in the chinook salmon (Oncorhynchus tshawytscha) caused by Phoma herbarum

Phoma herbarum has been associated with two outbreaks of systemic mycosis in hatchery-reared chinook salmon (Oncorhynchus tshawytscha) fingerlings. Affected fish exhibited abnormal swimming behavior, exophthalmia, multiple rounded areas of muscle softening, protruded hemorrhagic vents, and abdominal swelling. In all affected fish, swimbladders were filled with whitish creamy viscous fungal mass, surrounded by dark red areas in swimbladder walls, kidneys, and musculature. Clinical and histopathological examinations suggest that the infection may have started primarily in the swimbladder and then spread to the kidneys, gastrointestinal tract, and surrounding musculature. Consistent microscopical findings included broad septate branched fungal hyaline hyphae, 5–12 μm in diameter within the swimbladder, stomach, and often within and adjacent to blood vessels. Profuse growths of woolly brown fungal colonies were obtained from swimbladders and kidneys on Sabouraud medium. On corn meal agar the formation of pycnidia, characteristic of Phoma spp., was detected within 10 days of incubation. Morphological and molecular analyses identified this fungus as Phoma herbarum. This report underscores systemic fungal infections as a threat to raceway-raised salmon.     

Innate susceptibility differences in chinook salmon Oncorhynchus tshawytscha to Loma salmonae (Microsporidia)

Loma salmonae (Putz, Hoffman and Dunbar, 1965) Morrison & Sprague, 1981 (Microsporidia) is an important gill pathogen of Pacific salmon Oncorhynchus spp. in the Pacific Northwest. Three strains of chinook salmon O. tshawytscha were infected in 2 trials with L. salmonae by feeding of macerated infected gill tissue or per os as a gill tissue slurry. Intensity of infection was significantly higher in the Northern stream (NS) strain as compared to the Southern coastal (SC) and a hybrid (H) strain derived from these 2 strains. Both wet mount and histological enumeration of intensity of infection demonstrated strain differences. Survival in the NS strain was significantly lower than the other strains. The NS strain may represent a naive strain and be less able to mount an effective immune response against the parasite.     

Top-down and bottom-up control of life-history strategies in coho salmon (Oncorhynchus kisutch)

Sexual maturation profoundly affects population dynamics, but the degrees to which genetic, top-down, and bottom-up controls affect age at maturity are unclear. Salmonid fishes have plastic age at maturity, and we consider genetic and environmental effects on this trait by developing fitness functions for coho salmon (Oncorhynchus kisutch). The functions are based on size-specific survival and reproductive success, where reproductive success is the product of fecundity and ability to defend nests (females) or the product of sperm volume and ability to mate (males). We model genetic and bottom-up controls (e.g., food availability) with an environmentally explicit growth function and top-down control (predation mortality) with survival functions that consider both size-dependent and size-independent mortality. For females, we predict that early maturation rarely maximizes fitness, but males can maximize fitness by maturing early if they grow well in freshwater. We predict that early maturation is most affected by the bottom-up effects of resource distribution at sea, followed by bottom-up and genotypic effects in freshwater. Top-down processes are predicted to have strong effects on the likelihood of delayed maturation.     

Mortality of coho salmon (Oncorhynchus kisutch) associated with burdens of multiple parasite species

Multiple analytical techniques were used to evaluate the impact of multiple parasite species on the mortality of threatened juvenile coho salmon (Oncorhynchus kisutch) from the West Fork Smith River, Oregon, USA. We also proposed a novel parsimonious mathematical representation of macroparasite distribution, congestion rate, which (i) is easier to use than traditional models, and (ii) is based on Malthusian parameters rather than probability theory. Heavy infections of Myxobolus insidiosus (Myxozoa) and metacercariae of Nanophyetus salmincola and Apophallus sp. occurred in parr (subyearlings) from the lower mainstem of this river collected in 2007 and 2008. Smolts (yearlings) collected in 2007–2010 always harboured fewer Apophallus sp. with host mortality recognised as a function of intensity for this parasite. Mean intensity of Apophallus sp. in lower mainstem parr was 753 per fish in 2007 and 856 per fish in 2008, while parr from the tributaries had a mean of only 37 or 13 parasites per fish, respectively. Mean intensity of this parasite in smolts ranged between 47 and 251 parasites per fish. Over-dispersion (variance to mean ratios) of Apophallus sp. was always lower in smolts compared with all parr combined or lower mainstem parr. Retrospective analysis based on smolt data using both the traditional negative binomial truncation technique and our proposed congestion rate model showed identical results. The estimated threshold level for mortality involving Apophallus sp. was at 400–500 parasites per fish using both analytical methods. Unique to this study, we documented the actual existence of these heavy infections prior to the predicted mortality. Most of the lower mainstem parr (approximately 75%) had infections above this level. Heavy infections of Apophallus sp. metacercariae may be an important contributing factor to the high over-wintering mortality previously reported for these fish that grow and develop in this section of the river. Analyses using the same methods for M.insidiosus and N. salmincola generally pointed to minimal parasite-associated mortality.     

p,p'-DDE depresses the immune competence of chinook salmon (Oncorhynchus tshawytscha) leukocytes

p,p'-DDE, the main metabolite of DDT, is still detected in aquatic environments throughout the world. Here, the effects and mechanisms by which p,p'-DDE exposure might affect the immune system of chinook salmon (Oncorhynchus tshawytscha) was studied. Isolated salmon splenic and pronephric leukocytes were incubated with different concentrations of p,p'-DDE, and cell viability, induction of apoptosis, and mitogenic responses were measured by flow cytometry and Alamar Blue assay. p,p'-DDE significantly reduced cell viability and proliferation and increased apoptosis. The effect of p,p'-DDE on pronephric leukocytes was more severe than on splenic leukocytes, likely because pronephric leukocytes had a higher proportion of granulocytes, cells that appear more sensitive to p,p'-DDE. The effect of p,p'-DDE on leukocytes appeared to vary between developmental stages or seasonal differences. The mitogenic response of leukocytes of chinook salmon exposed to p,p'-DDE in vivo exhibited a biphasic dose-response relationship. Only leukocytes isolated from salmon treated with 59 ppm p,p'-DDE had a significantly lower percentage of Ig+ blasting cells than controls, although the response was biphasic. These results support the theory that exposure to chemical contaminants could lead to an increase in disease susceptibility and mortality of fish due to immune suppression.