Who are the missing parents? Grandparentage analysis identifies multiple sources of gene flow into a wild population

In order to increase the size of declining salmonid populations, supplementation programmes intentionally release fish raised in hatcheries into the wild. Because hatchery-born fish often have lower fitness than wild-born fish, estimating rates of gene flow from hatcheries into wild populations is essential for predicting the fitness cost to wild populations. Steelhead trout (Oncorhynchus mykiss) have both freshwater resident and anadromous (ocean-going) life history forms, known as rainbow trout and steelhead, respectively. Juvenile hatchery steelhead that 'residualize' (become residents rather than go to sea as intended) provide a previously unmeasured route for gene flow from hatchery into wild populations. We apply a combination of parentage and grandparentage methods to a three-generation pedigree of steelhead from the Hood River, Oregon, to identify the missing parents of anadromous fish. For fish with only one anadromous parent, 83% were identified as having a resident father while 17% were identified as having a resident mother. Additionally, we documented that resident hatchery males produced more offspring with wild anadromous females than with hatchery anadromous females. One explanation is the high fitness cost associated with matings between two hatchery fish. After accounting for all of the possible matings involving steelhead, we find that only 1% of steelhead genes come from residualized hatchery fish, while 20% of steelhead genes come from wild residents. A further 23% of anadromous steelhead genes come from matings between two resident parents. If these matings mirror the proportion of matings between residualized hatchery fish and anadromous partners, then closer to 40% of all steelhead genes come from wild trout each generation. These results suggest that wild resident fish contribute substantially to endangered steelhead 'populations' and highlight the need for conservation and management efforts to fully account for interconnected Oncorhynchus mykiss life histories.     

Genetic architecture of growth and early life‐history transitions in anadromous and derived freshwater populations of steelhead

Heritabilities of growth, precocious maturation and smolting were measured in 75 families of juvenile steelhead or rainbow trout Oncorhynchus mykiss , progeny of within and between line matings (crosses) of wild, anadromous steelhead and wild, resident (lake) rainbow trout originally derived from the same anadromous stock 70 years earlier. The tagged yearling progeny were combined by line in common freshwater rearing containers and graded into three categories: mature, smolt or rearing (undifferentiated) at age 2 years. Heritabilities of precocious male maturity, smolting and growth were moderate to high, and the genetic correlation between growth and smolting was low. Smolting and precocious male maturity were highly variable among families within lines and significantly different between lines. Each of the four lines produced significant numbers of smolts at age two. Smolting and maturation were negatively genetically correlated, which may explain the persistence of smolting in the lake population despite strong selection against lake smolts; balancing selection on male maturation age may help to maintain variation for smolting. The high heritability of smolting, coupled with the inability of smolts that leave the lake to return to it indicates that the genetic potential for smolting can lie dormant or be maintained through a dynamic interaction between smolting and early maturation for decades despite complete selection against the phenotype. The results have significant implications for the preservation of threatened anadromous stocks in fresh water and the inclusion of resident fish of formerly anadromous populations, currently trapped behind long‐standing barriers to migration, as one component of the same population.     

Discovery and characterization of single-nucleotide polymorphisms in steelhead/rainbow trout, Oncorhynchus mykiss

Single-nucleotide polymorphisms (SNPs) have several advantages over other genetic markers, including lower mutation and genotyping error rates, ease of inter-laboratory standardization, and the prospect of high-throughput, low-cost genotyping. Nevertheless, their development and use has only recently moved beyond model organisms to groups such as salmonid fishes. Oncorhynchus mykiss is a salmonid native to the North Pacific rim that has now been introduced throughout the world for fisheries and aquaculture. The anadromous form of the species is known as steelhead. Native steelhead populations on the west coast of the United States have declined and many now have protected status. The nonanadromous, or resident, form of the species is termed rainbow, redband or golden trout. Additional life history and morphological variation, and interactions between the forms, make the species challenging to study, monitor and evaluate. Here, we describe the discovery, characterization and assay development for 139 SNP loci in steelhead/rainbow trout. We used EST sequences from existing genomic databases to design primers for 480 genes. Sanger-sequencing products from these genes provided 130 KB of consensus sequence in which variation was surveyed for 22 individuals from steelhead, rainbow and redband trout groups. The resulting TaqMan assays were surveyed in five steelhead populations and three rainbow trout stocks, where they had a mean minor allele frequency of 0.15-0.26 and observed heterozygosity of 0.18-0.35. Mean F(ST) was 0.204. The development of SNPs for O. mykiss will help to provide highly informative genetic tools for individual and stock identification, pedigree reconstruction, phylogeography and ecological investigation.     

Evidence of partial anadromy and resident-form dispersal bias on a fine scale in populations of Oncorhynchus mykiss

We examine sympatric anadromous (steelhead) and nonanadromous (resident) rainbow trout (Oncorhynchus mykiss) from neighboring locations to test three hypotheses: (1) the sympatric life history types are not genetically different; (2) fine-scale dispersal is the same for both sexes, and (3) fine-scale dispersal is the same for steelhead and resident individuals. Data from 13 microsatellite loci reveal no genetic difference between sympatric steelhead and resident O. mykiss but moderate population structure (F _ST=0.019–0.028) between adjacent samples, regardless of life history type. Our results provide further evidence of partial anadromy and suggest that geographic proximity and genetic history, more than migratory type, should be considered when identifying populations for use in restoration of local genetic diversity. We find evidence of resident-form dispersal bias on a fine spatial scale, however, we find no evidence that fine-scale dispersal varies by gender. Conservation strategies should aim to maintain resident and anadromous forms when they occur in sympatry, as they may be important in facilitating gene flow on small and large spatial scales, respectively.     

Does light intensity affect self‐feeding and food wastage in group‐held rainbow trout and white‐spotted charr?

The self‐feeding rhythms of rainbow trout Oncorhynchus mykiss and white‐spotted charr Salvelinus leucomaenis were studied when group‐held fishes ( n  = 10 per group) were fed using self‐feeders under two different light intensities (50 lx, 16 μW cm⁻² and 700 lx, 215 μW cm⁻²) during the light phase of the light‐dark cycle. Food wastage was also measured. At 50 lx, all groups of rainbow trout learned to operate the self‐feeder within 4 days, whereas it took up to 25 days for all groups at 700 lx. In contrast, all groups of white‐spotted charr learned self‐feeding within 17 days, irrespective of light intensity. These results, although non‐significant, suggest that lower light intensities can stimulate instrumental learning in rainbow trout, but not white‐spotted charr. In rainbow trout, the total number of trigger actuations for the entire experimental period was significantly higher at 50 rather than 700 lx, although this may have been related to delayed learning at 700 lx. There was no significant effect in white‐spotted charr. Growth rate (assessed using the thermal growth coefficient) was also higher in rainbow trout but not white‐spotted charr at 50 rather than 700 lx, although this difference was non‐significant. Light intensity had no significant effect on food wastage in either rainbow trout or white‐spotted charr, and it did not appear to affect the proportion of trigger actuations during the light phase. Clear diurnal feeding rhythms were observed in both species and these were classified into four categories: uniform, dawn, dusk and crepuscular. At 50 lx, fish from both species generally fed in temporally localized periods at either dawn and dusk, whilst feeding was predominantly uniform during the light phase at 700 lx.     

Migration flexibility between freshwater and marine habitats of the pond smelt Hypomesus nipponensis

The life histories of the pond smelt Hypomesus nipponensis collected from Japanese fresh waters and brackish (sea) waters were studied by examining the strontium (Sr) and calcium (Ca) concentrations in their otoliths. The Sr:Ca ratios in the otoliths changed with the salinity of the habitat. The pond smelt living in a freshwater environment showed consistently low Sr:Ca ratios throughout the otolith, averaging 1·2–1·3 × 10⁻³. These samples were identified as a standard freshwater type. In contrast, fish collected from the intertidal zone showed higher otolith Sr:Ca ratios than those in the standard freshwater type, and the ratios fluctuated along the growth phase. In addition to the two representative life‐history types of H. nipponensis , i. e . freshwater and anadromous life‐history types, other pond smelts were found to have an estuarine resident life history‐type with no freshwater phase, indicating that the pond smelt has a flexible migration strategy with a high degree of behavioural plasticity and an ability to utilize the full range of salinity in its life history.     

Influence of temperature on silver accumulation and depuration in rainbow trout

To assess the influence of water temperature on silver uptake, rainbow trout Oncorhynchus mykiss ( c . 50 g; held at 13° C) were exposed to 0·1 μM AgNO₃ in ion‐poor water for 1 week at 4 and 16° C without previous temperature acclimation. To assess the influence of temperature on elimination of previously accumulated Ag, rainbow trout were exposed to 0·1 μM AgNO₃ in ion‐poor water for 1 week at 12° C, then were randomly divided amongst two Ag‐free water containers, differing only in temperature (3 and 16° C), for 2 months. In the uptake study greater accumulation of Ag was seen in the gills, plasma and especially the livers and bile of 'warm' rainbow trout (16° C) compared to 'cold' rainbow trout (4° C), which can be explained by the higher metabolic rates of the warmer fish. In the depuration study there was no net elimination of Ag from the livers and bile but there was biphasic elimination of Ag from the gills and plasma of 'warm' and 'cold' fish, but with few differences between them. This indicated that temperature‐dependent processes were less important in Ag elimination than in Ag uptake. Toxicokinetic modelling of Ag uptake by livers indicated four‐fold greater uptake of Ag by 'warm' rainbow trout compared to 'cold' rainbow trout (one compartment uptake model). Elimination of previously accumulated Ag from the plasma was best fitted by a two compartment rate‐constant based model, with approximately half the plasma Ag load eliminated within 24 h, followed by slower elimination of Ag over 2 months.     

Rapid parallel evolution of standing variation in a single,complex, genomic region is associated with life history in steelhead/rainbow trout

Rapid adaptation to novel environments may drive changes in genomic regions through natural selection. Such changes may be population-specific or, alternatively, may involve parallel evolution of the same genomic region in multiple populations, if that region contains genes or co-adapted gene complexes affecting the selected trait(s). Both quantitative and population genetic approaches have identified associations between specific genomic regions and the anadromous (steelhead) and resident (rainbow trout) life-history strategies of Oncorhynchus mykiss. Here, we use genotype data from 95 single nucleotide polymorphisms and show that the distribution of variation in a large region of one chromosome, Omy5, is strongly associated with life-history differentiation in multiple above-barrier populations of rainbow trout and their anadromous steelhead ancestors. The associated loci are in strong linkage disequilibrium, suggesting the presence of a chromosomal inversion or other rearrangement limiting recombination. These results provide the first evidence of a common genomic basis for life-history variation in O. mykiss in a geographically diverse set of populations and extend our knowledge of the heritable basis of rapid adaptation of complex traits in novel habitats.     

Enhancement of pH‐resistant iron‐binding activity in supernatants of rainbow trout blood leucocytes by in vitro treatment with phorbol‐12‐myristate‐13‐acetate

Leucocyte lysates from rainbow trout Oncorhynchus mykiss showed an iron‐binding activity that was retained even if the samples were exposed to an acid pH (4·5). Iron‐binding activity of leucocyte supernatants was enhanced by the presence of 1 μg ml⁻¹ phorbol‐12‐myristate‐13‐acetate in the cell medium.     

Regional variation in the microhardness and mineralization of vertebrae from brown and rainbow trout

Regional variation in properties of vertebral bone from brown Salmo trutta and rainbow trout Oncorhychus mykiss were explored by using microhardness tests. Statistically‐significant positive correlations were identified between the microhardness of bone and its mineral content. In both brown and rainbow trout, the vertebrae from the caudal region were harder than those of the trunk region. There was a significant difference between the species; microhardness of bone from vertebrae of rainbow trout was greater than those from brown trout.     

Gene Flow between Sympatric Life History Forms of Oncorhynchus mykiss Located above and below Migratory Barriers

Oncorhynchus mykiss have a diverse array of life history types, and understanding the relationship among types is important for management of the species. Patterns of gene flow between sympatric freshwater resident O. mykiss, commonly known as rainbow trout, and anadromous O. mykiss, commonly known as steelhead, populations are complex and poorly understood. In this study, we attempt to determine the occurrence and pathways of gene flow and the degree of genetic similarity between sympatric resident and anadromous O. mykiss in three river systems, and investigate whether resident O. mykiss are producing anadromous offspring in these rivers, two of which have complete barriers to upstream migration. We found that the population structure of the O. mykiss in these rivers appears to be influenced more by the presence of a barrier to upstream migration than by life history type. The sex ratio of resident O. mykiss located above a barrier, and smolts captured in screw traps was significantly skewed in favor of females, whereas the reverse was true below the barriers, suggesting that male resident O. mykiss readily migrate downstream over the barrier, and that precocious male maturation may be occurring in the anadromous populations. Through paternity analyses, we also provide direct confirmation that resident O. mykiss can produce offspring that become anadromous. Most (89%) of the resident O. mykiss that produced anadromous offspring were males. Our results add to the growing body of evidence that shows that gene flow does readily occur between sympatric resident and anadromous O. mykiss life history types, and indicates that resident O. mykiss populations may be a potential repository of genes for the anadromous life history type.     

Individual condition and stream temperature influence early maturation of rainbow and steelhead trout, Oncorhynchus mykiss

Alternative male phenotypes in salmonine fishes arise from individuals that mature as larger and older anadromous marine-migrants or as smaller and younger freshwater residents. To better understand the processes influencing the expression of these phenotypes we examined the influences of growth in length (fork length) and whole body lipid content in rainbow trout (Oncorhynchus mykiss). Fish were sampled from the John Day River basin in northeast Oregon where both anadromous (“steelhead”) and freshwater resident rainbow trout coexist. Larger males with higher lipid levels had a greater probability of maturing as a resident at age-1+. Among males, 38% were maturing overall, and the odds ratios of the logistic model indicated that the probability of a male maturing early as a resident at age-1+ increased 49% (95% confidence interval (CI) = 23–81%) for every 5 mm increase in length and 33% (95% CI = 10–61%) for every 0.5% increase in whole body lipid content. There was an inverse association between individual condition and water temperature as growth was greater in warmer streams while whole body lipid content was higher in cooler streams. Our results support predictions from life history theory and further suggest that relationships between individual condition, maturation, and environmental variables (e.g., water temperature) are shaped by complex developmental and evolutionary influences.     

Retention of a chromosomal inversion from an anadromous ancestor provides the genetic basis for alternative freshwater ecotypes in rainbow trout

Migratory behaviour patterns in animals are controlled by a complex genetic architecture. Rainbow trout (Oncorhynchus mykiss) is a salmonid fish that spawns in streams but exhibits three primary life history pathways: stream‐resident (fluvial), lake‐migrant (adfluvial) and ocean‐migrant (anadromous). Previous studies examining fluvial and anadromous O. mykiss have identified several genes associated with life history divergence including the presence of an inversion complex within chromosome 5 (Omy05) that appears to maintain a suite of linked genes controlling migratory behaviour. However, adfluvial trout are migratory without being anadromous, and the genetic basis for this life history has not been investigated from evolutionary perspectives. We sampled wild, native nonanadromous rainbow trout occupying connected stream and lake habitats in a southwest Alaskan watershed to determine whether these fish exhibit genetic divergence between fluvial and adfluvial ecotypes, and whether that divergence parallels that documented in fluvial and anadromous O. mykiss. Data from restriction site‐associated DNA (RAD) sequencing revealed an association between frequencies of both the Omy05 inversion complex and other single nucleotide polymorphisms (SNPs) with habitat type (stream or lake), supporting the genetic divergence of fluvial and adfluvial individuals in sympatry. The presence of a genetic basis for migration into lakes, analogous to that documented for anadromy, indicates that the adfluvial ecotype must be recognized separately from the fluvial form of O. mykiss even though neither is anadromous. These results highlight the genetic architecture underlying migration and the importance of chromosomal inversions in promoting and sustaining intraspecific diversity.     

Migration of hatchery‐reared Atlantic salmon and wild anadromous brown trout post‐smolts in a Norwegian fjord system

Hatchery‐reared Atlantic salmon Salmo salar ( n  = 25) and wild anadromous brown trout (sea trout) Salmo trutta ( n  = 15) smolts were tagged with coded acoustic transmitters and released at the mouth of the River Eira on the west coast of Norway. Data logging receivers recorded the fish during their outward migration at 9, 32, 48 and 77 km from the release site. Seventeen Atlantic salmon (68%) and eight sea trout (53%) were recorded after release. Mean migratory speeds between different receiver sites ranged from 0·49 to 1·82 body lengths (total length) per second (bl s⁻¹) for Atlantic salmon and 0·11–2·60 bl s⁻¹ for sea trout. Atlantic salmon were recorded 9, 48 and 77 km from the river mouth on average 28, 65 and 83 h after release, respectively. Sea trout were recorded 9 km from the release site 438 h after release. Only four (23%) sea trout were detected in the outer part of the fjord system, while the rest of the fish seemed to stay in the inner fjord system. The Atlantic salmon stayed for a longer time in the inner part than in the outer parts of the fjord system, but distinct from sea trout, migrated through the whole fjord system into the ocean.     

Asymmetry in sexual development of gonads in intersex rainbow trout

Large‐scale sampling of spontaneous rainbow trout Oncorhynchus mykiss intersexes indicated a strong asymmetry of gonad differentiation in XX females; the right gonad was more sensitive to the mutation‐induced masculinization than the left one.     

Constraints of body size and swimming velocity on the ability of juvenile rainbow trout to endure periods without food

The hypothesis that body size and swimming velocity affect proximate body composition, wet mass and size‐selective mortality of fasted fish was evaluated using small (107 mm mean total length, L _T) and medium (168 mm mean L _T) juvenile rainbow trout Oncorhynchus mykiss that were sedentary or swimming ( c . 1 or 2 body length s⁻¹) and fasted for 147 days. The initial amount of energy reserves in the bodies of fish varied with L _T. Initially having less lipid mass and relatively higher mass‐specific metabolic rates caused small rainbow trout that were sedentary to die of starvation sooner and more frequently than medium‐length fish that were sedentary. Swimming at 2 body length s⁻¹ slightly increased the rate of lipid catabolism relative to 1 body length s⁻¹, but did not increase the occurrence of mortality among medium fish. Death from starvation occurred when fish had <3·2% lipid remaining in their bodies. Juvenile rainbow trout endured long periods without food, but their ability to resist death from starvation was limited by their length and initial lipid reserves.     

Signatures of natural selection among lineages and habitats in Oncorhynchus mykiss

Recent advances in molecular interrogation techniques now allow unprecedented genomic inference about the role of adaptive genetic divergence in wild populations. We used high-throughput genotyping to screen a genome-wide panel of 276 single nucleotide polymorphisms (SNPs) for the economically and culturally important salmonid Oncorhynchus mykiss. Samples included 805 individuals from 11 anadromous and resident populations from the northwestern United States and British Columbia, and represented two major lineages including paired populations of each life history within single drainages of each lineage. Overall patterns of variation affirmed clear distinctions between lineages and in most instances, isolation by distance within them. Evidence for divergent selection at eight candidate loci included significant landscape correlations, particularly with temperature. High diversity of two nonsynonymous mutations within the peptide-binding region of the major histocompatibility complex (MHC) class II (DAB) gene provided signatures of balancing selection. Weak signals for potential selection between sympatric resident and anadromous populations were revealed from genome scans and allele frequency comparisons. Our results suggest an important adaptive role for immune-related functions and present a large genomic resource for future studies.     

Fluvial rainbow trout contribute to the colonization of steelhead (Oncorhynchus mykiss) in a small stream

Life history polymorphisms provide ecological and genetic diversity important to the long term persistence of species responding to stochastic environments. Oncorhynchus mykiss have complex and overlapping life history strategies that are also sympatric with hatchery populations. Passive integrated transponder (PIT) tags and parentage analysis were used to identify the life history, origin (hatchery or wild) and reproductive success of migratory rainbow/steelhead for two brood years after barriers were removed from a small stream. The fluvial rainbow trout provided a source of wild genotypes to the colonizing population boosting the number of successful spawners. Significantly more parr offspring were produced by anadromous parents than expected in brood year 2005, whereas significantly more parr offspring were produced by fluvial parents than expected in brood year 2006. Although hatchery steelhead were prevalent in the Methow Basin, they produced only 2 parr and no returning adults in Beaver Creek. On average, individual wild steelhead produced more parr offspring than the fluvial or hatchery groups. Yet, the offspring that returned as adult steelhead were from parents that produced few parr offspring, indicating that high production of parr offspring may not be related to greater returns of adult offspring. These data in combination with other studies of sympatric life histories of O. mykiss indicate that fluvial rainbow trout are important to the conservation and recovery of steelhead and should be included in the management and recovery efforts.     

不同倍性虹鳟幼鱼对急性温度胁迫的抗氧化响应

虹鳟属于冷水性鱼类,其生存的最适温度为12~18 ℃,温度胁迫是虹鳟在夏秋季养殖过程中经常遇到的问题.枫叶鲑和硬头鳟均为品质优良的虹鳟选育种.为探讨急性温度胁迫对两种倍性虹鳟抗氧化响应的影响,本研究选取二倍体枫叶鲑和三倍体硬头鳟幼鱼,分别在13、17、21和25 ℃下进行热应激试验.在达到目标温度后的0、1、6和12 h取样,之后将受试鱼恢复至13 ℃的适宜温度下培养,并在恢复培养的1、12、24和48 h后取样,测定受试鱼肝脏中超氧化物歧化酶(SOD)、过氧化氢酶(CAT)、谷胱甘肽过氧化物酶(GPx)的活性,以及脂质过氧化物(LPO)、丙二醛(MDA)和热激蛋白70(HSP70)含量.结果表明: 枫叶鲑和硬头鳟的3种抗氧化酶活性在17 ℃组没有出现显著升高;21 ℃组枫叶鲑和硬头鳟的SOD活性在热应激期间出现显著升高,但枫叶鲑的SOD活性在恢复过程中恢复到正常水平;25 ℃组枫叶鲑和硬头鳟的SOD、CAT和GPx的活性均显著提高;但在恢复试验进行24 h后,CAT和GPx的活性恢复到正常水平.枫叶鲑在17、21和25 ℃组HSP70的产生量显著高于13 ℃组,而硬头鳟仅在21和25 ℃组HSP70的产生量显著高于13 ℃组.通过整合生物标志物响应指标值对多种抗氧化参数进行分析,发现枫叶鲑在17和21 ℃组急性温度胁迫下的抗氧化响应显著高于硬头鳟,但在25 ℃组硬头鳟的抗氧化响应高于枫叶鲑.这表明不同倍性虹鳟幼鱼在不同温度急性胁迫下的抗氧化响应有所差异.     

Genetic relationships among populations of wild resident, and wild and hatchery anadromous brook charr

To determine the genetic relationship of anadromous and resident life-history types within and among drainages, and compare several hatchery strains to their progenitor populations, brook charr Salvelinus fontinalis were examined for allozyme and mitochondrial DNA variation. Greater genetic similarity of sympatric anadromous and resident charr was found compared to similar life-history forms allopatrically, suggesting the two life-history types are not reproductively isolated. Low divergence among the mtDNA haplotypes suggests that the two life-history types are members of the same evolutionary lineage. Population differentiation from mtDNA data exceeded that from estimates based on allozymes. Genetic deviations from expectations suggest that the hatchery strains were derived from few individuals.