Multigenerational outbreeding effects in Chinook salmon (Oncorhynchus tshawytscha)

Outbreeding, mating between genetically divergent individuals, may result in negative fitness consequences for offspring via outbreeding depression. Outbreeding effects are of notable concern in salmonid research as outbreeding can have major implications for salmon aquaculture and conservation management. We therefore quantified outbreeding effects in two generations (F₁ hybrids and F₂ backcrossed hybrids) of Chinook salmon (Oncorhynchus tshawytscha) derived from captively-reared purebred lines that had been selectively bred for differential performance based on disease resistance and growth rate. Parental lines were crossed in 2009 to create purebred and reciprocal hybrid crosses (n = 53 families), and in 2010 parental and hybrid crosses were crossed to create purebred and backcrossed hybrid crosses (n = 66 families). Although we found significant genetic divergence between the parental lines (F_ST = 0.130), reciprocal F₁ hybrids showed no evidence of outbreeding depression (hybrid breakdown) or favorable heterosis for weight, length, condition or survival. The F₂ backcrossed hybrids showed no outbreeding depression for a suite of fitness related traits measured from egg to sexually mature adult life stages. Our study contributes to the current knowledge of outbreeding effects in salmonids and supports the need for more research to better comprehend the mechanisms driving outbreeding depression.     

The complete sequence of the mitochondrial genome of the Chinook salmon, Oncorhynchus tshawytscha

The complete sequence of the mitochondrial genome of Chinook salmon, Oncorhynchus tshawytscha, has been determined. The circular genome consisting of 16,644 base pairs encodes thirteen proteins, the 12S and 16S ribosomal RNAs, and 22 transfer RNAs. These genes are ordered in the same way as most other vertebrates. The nucleotide and amino acid sequences of the ribosomal RNAs and the thirteen protein-coding genes were compared with those of other salmonids such as Oncorhynchus mykiss, Salmo salar, Salvelinus fontinalis, Salvelinus alpinus and Coregonus lavaretus. The sequence features of the control region (D-loop), the origin of L-strand replication and a putative peptide codified by the 16S mitochondrial RNA are described and discussed.     

Population differences in Chinook salmon (Oncorhynchus tshawytscha) DNA methylation: Genetic drift and environmental factors

Local adaptation and phenotypic differences among populations have been reported in many species, though most studies focus on either neutral or adaptive genetic differentiation. With the discovery of DNA methylation, questions have arisen about its contribution to individual variation in and among natural populations. Previous studies have identified differences in methylation among populations of organisms, although most to date have been in plants and model animal species. Here we obtained eyed eggs from eight populations of Chinook salmon (Oncorhynchus tshawytscha) and assayed DNA methylation at 23 genes involved in development, immune function, stress response, and metabolism using a gene‐targeted PCR‐based assay for next‐generation sequencing. Evidence for population differences in methylation was found at eight out of 23 gene loci after controlling for developmental timing in each individual. However, we found no correlation between freshwater environmental parameters and methylation variation among populations at those eight genes. A weak correlation was identified between pairwise DNA methylation dissimilarity among populations and pairwise F _ST based on 15 microsatellite loci, indicating weak effects of genetic drift or geographic distance on methylation. The weak correlation was primarily driven by two genes, GTIIBS and Nkef. However, single‐gene Mantel tests comparing methylation and pairwise F _ST were not significant after Bonferroni correction. Thus, population differences in DNA methylation are more likely related to unmeasured oceanic environmental conditions, local adaptation, and/or genetic drift. DNA methylation is an additional mechanism that contributes to among population variation, with potential influences on organism phenotype, adaptive potential, and population resilience.     

Gauging resource exploitation by juvenile Chinook salmon (Oncorhynchus tshawytscha) in restoring estuarine habitat

In the context of delta restoration and its impact on salmonid rearing, success is best evaluated based on whether out‐migrating juvenile salmon can access and benefit from suitable estuarine habitat. Here, we integrated 3 years of post‐restoration monitoring data including habitat availability, invertebrate prey biomass, and juvenile Chinook salmon (Oncorhynchus tshawytscha) physiological condition to determine whether individuals profited from the addition of 364 ha of delta habitat in South Puget Sound, Washington, United States. Productivity in the restored mudflat was comparable to reference sites 3 years after dike removal, surpassing a mean total of 6 million kJ energy from invertebrate prey. This resulted from the development of a complex network of tidal channels and a resurgence in dipteran biomass that was unique to the restoration area. Consequently, a notable shift in invertebrate consumption occurred between 2010 and 2011, whereby individuals switched from eating primarily amphipods to dipteran flies; however, dietary similarity to the surrounding habitat did not change from year to year, suggesting that this shift was a result of a change in the surrounding prey communities. Growth rates did not differ between restored and reference sites, but catch weight was positively correlated with prey biomass, where greater prey productivity appeared to offset potential density‐dependent effects. These results demonstrate how the realized function of restoring estuarine habitat is functionally dependent. High prey productivity in areas with greater connectivity may support healthy juvenile salmon that are more likely to reach the critical size class for offshore survival.     

Behavioural thermoregulation by subyearling fall (autumn) Chinook salmon Oncorhynchus tshawytscha in a reservoir

This study investigated behavioural thermoregulation by subyearling fall (autumn) Chinook salmon Oncorhynchus tshawytscha in a reservoir on the Snake River, Washington, U.S.A. During the summer, temperatures in the reservoir varied from 23° C on the surface to 11° C at 14 m depth. Subyearlings implanted with temperature-sensing radio transmitters were released at the surface at temperatures >20° C during three blocks of time in summer 2004. Vertical profiles were taken to measure temperature and depth use as the fish moved downstream over an average of 5·6–7·2 h and 6·0–13·8 km. The majority of the subyearlings maintained average body temperatures that differed from average vertical profile temperatures during most of the time they were tracked. The mean proportion of the time subyearlings tracked within the 16–20° C temperature range was larger than the proportion of time this range was available, which confirmed temperature selection opposed to random use. The subyearlings selected a depth and temperature combination that allowed them to increase their exposure to temperatures of 16–20° C when temperatures <16 and >20° C were available at lower and higher positions in the water column. A portion of the subyearlings that selected a temperature c. 17·0° C during the day, moved into warmer water at night coincident with an increase in downstream movement rate. Though subyearlings used temperatures outside of the 16–20° C range part of the time, behavioural thermoregulation probably reduced the effects of intermittent exposure to suboptimal temperatures. By doing so, it might enhance growth opportunity and life-history diversity in the population of subyearlings studied.     

The disulfide bond pattern of salmon egg lectin 24K from the Chinook salmon Oncorhynchus tshawytscha

The disulfide bonds in the galactose-specific lectin SEL 24K from the egg of the Chinook salmon Oncorhynchus tshawytscha were determined by mass spectrometry. Four predictive in silico tools were used to determine the oxidation state of cysteines in the sequence and possible location of the disulfide bonds. A combination of tryptic digestion, HPLC separation, and chemical modifications were used to establish the location of seven disulfide bonds and one pair of free cysteines. After proteolysis, peptides containing one or two disulfide bonds were identified by reduction and mass spectral comparison. MALDI mass spectrometry was supported by chemical modification (iodoacetamide) and in silico digestion. The assignments of disulfide bonds were further confirmed by mass spectral fragmentation studies including in-source dissociation (ISD) and collision-induced dissociation (CID). The experimentally determined disulfide bonds and free Cys residues were only partially consistent with those generated by several automated public-domain algorithms.     

Environmental adaptation in Chinook salmon (Oncorhynchus tshawytscha) throughout their North American range

Landscape genomics is a rapidly growing field with recent advances in both genotyping efficiency and statistical analyses that provide insight towards local adaptation of populations under varying environmental and selective pressure. Chinook salmon (Oncorhynchus tshawytscha) are a broadly distributed Pacific salmon species, occupying a diversity of habitats throughout the northeastern Pacific with pronounced variation in environmental and climate features but little is understood regarding local adaptation in this species. We used a multivariate method, redundancy analysis (RDA), to identify polygenic correlations between 19 703 SNP loci and a suite of environmental variables in 46 collections of Chinook salmon (1956 total individuals) distributed throughout much of its North American range. Models in RDA were conducted on both rangewide and regional scales by hierarchical partitioning of the populations into three distinct genetic lineages. Our results indicate that between 5.8 and 21.8% of genomic variation can be accounted for by environmental features, and 566 putatively adaptive loci were identified as targets of environmental adaptation. The most influential drivers of adaptive divergence included precipitation in the driest quarter of the year (Rangewide and North Coastal Lineage, anova P = 0.002 and 0.01, respectively), precipitation in the wettest quarter of the year (Interior Columbia River Stream‐Type Lineage, anova P = 0.03), variation in mean diurnal range in temperature (South Coastal Lineage, anova P = 0.005), and migration distance (Rangewide, anova P = 0.001). Our results indicate that environmental features are strong drivers of adaptive genomic divergence in this species, and provide a foundation to investigate how Chinook salmon might respond to global environmental change.     

Variation of mitochondrial DNA in Chinook salmon Oncorhynchus tschawytscha Walbaum populations from Kamchatka

The variation in mitochondrial DNA (mtDNA) structure among Chinook Salmon Oncorhynchus tschawytscha Walbaum populations from Kamchatka was inferred from restriction length polymorphism analysis using eight restriction endonucleases. The nucleotide sequence variation in three amplified mtDNA regions was examined at seven polymorphic restriction sites in 579 fish from 13 localities. Based on the frequencies of 11 combined haplotypes and the number of nucleotide substitutions, the among- and within-population variation was estimated. The heterogeneity test showed highly significant differences among all the populations. The estimated maximum time of independent divergence of the Asian Chinook salmon populations, whose differences was about 0.02% nucleotide substitutions, did not exceed 10000-20000 years. Apparently, the retreat of the late Pleistocene glacier triggered spreading, recolonization, and formation of the present-day pattern of the species subdivision into structural components.     

Spatial and temporal patterns of growth and consumption by juvenile spring/summer Chinook salmon Oncorhynchus tshawytscha

Somatic growth is often used as a metric of habitat quality, but such an approach has limitations because growth results from complex interactions between abiotic and biotic factors. In this study, we derived estimates of weekly growth (based on otolith growth increments) across several months for four populations of threatened Chinook salmon, Oncorhynchus tshawytscha, from the Salmon River Basin, Idaho, USA. Although mean stream temperature varied by 2–5 °C across populations, growth across the season did not vary significantly by population. To investigate this further, we applied a bioenergetics model that produced estimates of consumption rates. We then examined how growth and consumption rates varied according to rearing stream and day in the season. Using generalized least squares models, somatic growth (g?day^?1) was best explained by stream and date, yet a model with only date had moderate support, and thus indicated limited support for stream effects. Specific consumption rate (g?g^?1?day^?1 and J?g^?1?day^?1) was best explained using a model that included main effects of stream and date. These findings are consistent with the hypothesis that higher temperatures confer higher metabolic costs that require greater consumption to produce similar growth rates in cooler streams. This highlights that similarity in growth rate among streams may mask changes in individual behavior and/or energetic acquisition associated with differences in temperatures among streams. Results of this study represent the first steps towards identifying factors that underlie important population level and habitat quality differences.     

Cardiovascular responses of Chinook salmon (Oncorhynchus tshawytscha) during rapid anaesthetic induction and recovery

The effects of three anaesthetics on induction and recovery were compared in Chinook salmon (Oncorhynchus tshawytscha). Heart rate (HR), cardiac output (Q), dorsal aortic pressure (DAP) and stroke volume (SV) were measured in minimally disturbed salmon during 5 min anaesthetic inductions with approximately equi-potent concentrations of MS222 (100 ppm), metomidate (6-10 ppm) and AQUI-S (60 ppm). MS222 induction caused a steady decline in DAP only, while metomidate induction did not affect any cardiovascular variable. AQUI-S caused a biphasic response, and within 2 min had depressed HR, Q, DAP and SV by between 20 and 50%. In the final 3 min HR returned to pre-anaesthesia levels, and Q and SV climbed to greater than pre-anaesthesia levels. Blood samples taken pre- and post-anaesthesia showed all inductions caused hypoxaemia (oxygen partial pressure of dorsal aortic blood (PaO2): MS222 47 mmHg, metomidate 35 mmHg, AQUI-S 21 mmHg). Haematocrit and plasma adrenaline and noradrenaline levels increased slightly in AQUI-S treated fish only. Recovery was monitored for 6 h post-anaesthesia, and was similar for each anaesthetic. All cardiovascular variables had returned to control levels within 5 min with the exception of DAP, which was initially slightly elevated (up to 20%) but returned to control values within 30 min. Anaesthesia is usually preceded by handling. Netting prior to anaesthesia caused significant increases in HR, Q and SV, which masked any anaesthetic dependent effects. Recovery from anaesthesia combined with surgery was also generally anaesthetic independent and recovery was prolonged, compared to anaesthesia alone. These data suggest limiting fish handling/manipulation is more important in minimising cardiovascular disturbance than the choice of anaesthetic.     

Juvenile growth and aggression in diploid and triploid Chinook salmon Oncorhynchus tshawytscha (Walbaum)

Multilocus heterozygosity, aggressive and feeding behaviour, plasma cortisol levels and growth rate were evaluated among three groups of juvenile Chinook salmon Oncorhynchus tshawytscha : diploid, triploid and mixed groups of diploid and triploid fish. There was no difference between diploid and triploid fish in measurements of heterozygosity calculated using seven microsatellite loci, and these measurements did not correlate with performance measurements including feeding rate and growth rate. Aggression trials that examined small groups of fish revealed that after 4 days together in tanks, triploid fish were significantly less aggressive during feeding than diploid fish or fish in mixed groups. At the end of the trials, however, plasma cortisol levels did not differ among the three groups. Thirty-day growth trials in duplicate tanks of 60 fish revealed no difference in growth rate among diploid, triploid and mixed groups, but plasma cortisol levels were significantly lower in triploid fish than in either diploid fish or the mixed fish. Overall, independent of the above differences in aggressive behaviour and cortisol levels, these results suggest similar performance in diploid and triploid Chinook salmon, and thus provide support for the viability of triploid Chinook salmon culture in commercial aquaculture.     

The effects of environmental factors on the migratory movement patterns of Sacramento River yearling late-fall run Chinook salmon (Oncorhynchus tshawytscha)

Understanding smolt migration dynamics is a critical step in the preservation and conservation of imperiled salmonids in California’s Sacramento River system. Late-fall run Chinook salmon yearling smolts were acoustically tagged and tracked during their outmigration through California’s Sacramento River and San Francisco Estuary during 2007–2009. Migration rates were 14.3 km·day^-1 (± 1.3 S.E.) to 23.5 km?day^-1 (± 3.6 S.E.), similar to rates published for other West Coast yearling Chinook salmon smolt emigrations. Region-specific movement rates were fastest through the upper river regions, and slowest in the Sacramento/San Joaquin River Delta. River travel times were recorded for smolts travelling through a series of ten monitor-delimited reaches. Using these, a smolt travel time model determined by two parameters (movement rate and rate of population spreading) was then used to determine the influence of different factors on the model’s fit, using model selection with Akaike’s Information Criterion. The model that allowed for both year and reach to be expressed additively for both travel time and population spreading rate estimates, while accounting for a “release” effect, was the best supported model. Finally, several models incorporated environmental data as a linear predictor of movement rates. The addition of the environmental variables, in order of importance, river width to depth ratio, river flow, water turbidity, river flow to mean river flow ratio, and water velocity all resulted in improved model fit. Water temperature did not improve model fit. These environmental associations are discussed and potential improvements on the travel time model are suggested.     

Captive rearing for Chinook salmon (Oncorhynchus tshawytscha) and Atlantic salmon (Salmo salar): the Idaho and Maine experiences

Captive rearing is a conservation strategy where juveniles are collected from the natural environment, reared to maturity in a hatchery environment, and then released back into the natural environment at maturity for volitional spawning. This strategy has been used to produce adult outplants for stock enhancement where natural escapement is poor or capture of adults is difficult. In both Idaho (Chinook salmon, Oncorhynchus tshawytscha) and Maine (Atlantic salmon, Salmo salar), captive rearing programs have been initiated as an experimental strategy to prevent cohort collapse and conserve genetic integrity of select depressed populations. In this paper, we provide an overview of these programs and describe some of the methods used to evaluate the effectiveness of this approach. Behaviors such as habitat selection, courting, and spawn timing were monitored. Data collected for both programs indicate that the captive fish display similar behaviors as their wild conspecifics in terms of habitat selection and spawning, although there were some differences in spawn timing. Evaluations of egg and fry production also indicate that captive-reared adults are successfully spawning and producing offspring. Each program is still waiting on final evaluations of reproductive success through genetic analyses of returning adults, but results so far indicate that this could be an additional captive propagation strategy for depressed populations.     

Tactic‐specific benefits of polyandry in Chinook salmon Oncorhynchus tshawytscha

This study examined whether polyandrous female Chinook salmon Oncorhynchus tshawytscha obtain benefits compared with monandrous females through an increase in hatching success. Both of the alternative reproductive tactics present in male O. tshawytscha (large hooknoses and small, precocious jacks) were used, such that eggs were either fertilized by a single male (from each tactic) or multiple males (using two males from the same or different tactics). The results show that fertilized eggs from the polyandrous treatments had a significantly higher hatching success than those from the monandrous treatments. It is also shown that sperm speed was positively related with offspring hatching success. Finally, there were tactic‐specific effects on the benefits females received. The inclusion of jacks in any cross resulted in offspring with higher hatching success, with the cross that involved a male from each tactic providing offspring with the highest hatching success than any other cross. This study has important implications for the evolution of multiple mating and why it is so prevalent across taxa, while also providing knowledge on the evolution of mating systems, specifically those with alternative reproductive tactics.     

Candidate loci reveal genetic differentiation between temporally divergent migratory runs of Chinook salmon (Oncorhynchus tshawytscha)

Local adaptation is a dynamic process driven by selection that can vary both in space and time. One important temporal adaptation for migratory animals is the time at which individuals return to breeding sites. Chinook salmon (Oncorhynchus tshawytscha) are excellent subjects for studying the genetic basis of temporal adaptation because their high seasonal homing fidelity promotes reproductive isolation leading to the formation of local populations across diverse environments. We tested for adaptive genetic differentiation between seasonal runs of Chinook salmon using two candidate loci; the circadian rhythm gene, OtsClock1b, and Ots515NWFSC, a microsatellite locus showing sequence identity to three salmonid genes central to reproductive development. We found significant evidence for two genetically distinct migratory runs in the Feather River, California (OtsClock1b: F(ST)=0.042, P=0.02; Ots515NWFSC: F(ST)=0.058, P=0.003). In contrast, the fall and threatened spring runs are genetically homogenous based on neutral microsatellite data (F(ST)=-0.0002). Similarly, two temporally divergent migratory runs of Chinook salmon from New Zealand are genetically differentiated based on polymorphisms in the candidate loci (OtsClock1b: F(ST)=0.083, P-value=0.001; Ots515NWFSC: F(ST)=0.095, P-value=0.000). We used an individual-based assignment method to confirm that these recently diverged populations originated from a single source in California. Tests for selective neutrality indicate that OtsClock1b and Ots515NWFSC exhibit substantial departures from neutral expectations in both systems. The large F(ST )estimates could therefore be the result of directional selection. Evidence presented here suggests that OtsClock1b and Ots515NWFSC may influence migration and spawning timing of Chinook salmon in these river systems.     

Proteomic Analysis of Chinook Salmon (Oncorhynchus tshawytscha) Ovarian Fluid

The ovarian, or coelomic, fluid that is released with the egg mass of many fishes is increasingly found to play an important role in several biological processes crucial for reproductive success. These include maintenance of oocyte fertility and developmental competence, prolonging of sperm motility, and enhancing sperm swimming speed. Here we examined if and how the proteome of chinook salmon (Oncorhynchus tshawytscha) ovarian fluid varied among females and then sought to examine the composition of this fluid. Ovarian fluid in chinook salmon was analyzed using 1D SDS PAGE and LC-MS/MS tryptic digest screened against Mascot and Sequest databases. We found marked differences in the number and concentrations of proteins in salmon ovarian fluid across different females. A total of 174 proteins were identified in ovarian fluid, 47 of which were represented by six or more peptides, belonging to one of six Gene Ontology pathways. The response to chemical stimulus and response to hypoxia pathways were best represented, accounting for 26 of the 174 proteins. The current data set provides a resource that furthers our understanding of those factors that influence successful egg production and fertilisation in salmonids and other species.     

An integrated linkage map reveals candidate genes underlying adaptive variation in Chinook salmon (Oncorhynchus tshawytscha)

Salmonids are an important cultural and ecological resource exhibiting near worldwide distribution between their native and introduced range. Previous research has generated linkage maps and genomic resources for several species as well as genome assemblies for two species. We first leveraged improvements in mapping and genotyping methods to create a dense linkage map for Chinook salmon Oncorhynchus tshawytscha by assembling family data from different sources. We successfully mapped 14 620 SNP loci including 2336 paralogs in subtelomeric regions. This improved map was then used as a foundation to integrate genomic resources for gene annotation and population genomic analyses. We anchored a total of 286 scaffolds from the Atlantic salmon genome to the linkage map to provide a framework for the placement 11 728 Chinook salmon ESTs. Previously identified thermotolerance QTL were found to colocalize with several candidate genes including HSP70, a gene known to be involved in thermal response, as well as its inhibitor. Multiple regions of the genome with elevated divergence between populations were also identified, and annotation of ESTs in these regions identified candidate genes for fitness related traits such as stress response, growth and behaviour. Collectively, these results demonstrate the utility of combining genomic resources with linkage maps to enhance evolutionary inferences.     

Using parentage analysis to estimate rates of straying and homing in Chinook salmon (Oncorhynchus tshawytscha)

We used parentage analysis based on microsatellite genotypes to measure rates of homing and straying of Chinook salmon (Oncorhynchus tshawytscha) among five major spawning tributaries within the Wenatchee River, Washington. On the basis of analysis of 2248 natural‐origin and 11594 hatchery‐origin fish, we estimated that the rate of homing to natal tributaries by natural‐origin fish ranged from 0% to 99% depending on the tributary. Hatchery‐origin fish released in one of the five tributaries homed to that tributary at a far lower rate than the natural‐origin fish (71% compared to 96%). For hatchery‐released fish, stray rates based on parentage analysis were consistent with rates estimated using physical tag recoveries. Stray rates among major spawning tributaries were generally higher than stray rates of tagged fish to areas outside of the Wenatchee River watershed. Within the Wenatchee watershed, rates of straying by natural‐origin fish were significantly affected by spawning tributary and by parental origin: progeny of naturally spawning hatchery‐produced fish strayed at significantly higher rates than progeny whose parents were themselves of natural origin. Notably, none of the 170 offspring that were products of mating by two natural‐origin fish strayed from their natal tributary. Indirect estimates of gene flow based on F_ST statistics were correlated with but higher than the estimates from the parentage data. Tributary‐specific estimates of effective population size were also correlated with the number of spawners in each tributary.