Social environment affects sensory gene expression in ant larvae.

Social insects depend on communication to regulate social behaviour. This also applies to their larvae, which are commonly exposed to social interactions and can react to social stimulation. However, how social insect larvae sense their environment is not known. Using RNAseq, we characterized expression of sensory-related genes in larvae of the ant Formica fusca, upon exposure to two social environments: isolation without contact to other individuals, and stimulation via the presence of other developing individuals. Expression of key sensory-related genes was higher following social stimulation, and larvae expressed many of the same sensory-related genes as adult ants and larvae of other insects, including genes belonging to the major insect chemosensory gene families. Our study provides first insights into the molecular changes associated with social information perception in social insect larvae.

Widespread hybridization within mound-building wood ants in Southern Finland results in cytonuclear mismatches and potential for sex-specific hybrid breakdown.

Hybridization and gene flow between diverging lineages are increasingly recognized as common evolutionary processes, and their consequences can vary from hybrid breakdown to adaptive introgression. We have previously found a population of wood ant hybrids between Formica aquilonia and F. polyctena that shows antagonistic effects of hybridization: females with introgressed alleles show hybrid vigour, whereas males with the same alleles show hybrid breakdown. Here, we investigate whether hybridization is a general phenomenon in this species pair and analyse 647 worker samples from 16 localities in Finland using microsatellite markers and a 1200-bp mitochondrial sequence. Our results show that 27 sampled nests contained parental-like gene pools (six putative F. polyctena and 21 putative F. aquilonia) and all remaining nests (69), from nine localities, contained hybrids of varying degrees. Patterns of genetic variation suggest these hybrids arise from several hybridization events or, instead, have backcrossed to the parental gene pools to varying extents. In contrast to expectations, the mitochondrial haplotypes of the parental species were not randomly distributed among the hybrids. Instead, nests that were closer to parental-like F. aquilonia for nuclear markers preferentially had F. polyctena's mitochondria and vice versa. This systematic pattern suggests there may be underlying selection favouring cytonuclear mismatch and hybridization. We also found a new hybrid locality with strong genetic differences between the sexes similar to those predicted under antagonistic selection on male and female hybrids. Further studies are needed to determine the selective forces that act on male and female genomes in these newly discovered hybrids.

Cardiac reflexes in a warming world: thermal plasticity of barostatic control and autonomic tones in a temperate fish.

Thermal plasticity of cardiorespiratory function allows ectotherms like fish to cope with seasonal temperature changes and is critical for resilience to climate change. Yet, the chronic thermal effects on cardiovascular homeostatic reflexes in fish are little understood although this may have important implications for physiological performance and overall resilience to climate warming. We compared cardiac autonomic control and baroreflex regulation of heart rate in perch (Perca fluviatilis L.) from a reference area in the Baltic Sea at 18-19°C with conspecifics from the Biotest enclosure, a chronically heated ecosystem receiving warmed effluent water (24-25°C) from a nuclear power plant. Resting heart rate of Biotest fish displayed clear thermal compensation and was 58.3±2.3 beats min-1 compared with 52.4±2.6 beats min-1 in reference fish at their respective environmental temperatures (Q10=1.2). The thermally compensated heart rate of Biotest fish was a combined effect of elevated inhibitory cholinergic tone (105% in Biotest fish versus 70% in reference fish) and reduced intrinsic cardiac pacemaker rate. A barostatic response was evident in both groups, as pharmacologically induced increases and decreases in blood pressure resulted in atropine-sensitive bradycardia and tachycardia, respectively. Yet, the tachycardia in Biotest fish was significantly greater, presumably due to the larger scope for vagal release. Acclimation of Biotest fish to 18°C for 3 weeks abolished differences in intrinsic heart rate and autonomic tone, suggesting considerable short-term thermal plasticity of cardiovascular control in this species. The heightened hypotensive tachycardia in Biotest perch may represent an important mechanism of ectothermic vertebrates that safeguards tissue perfusion pressure when tissue oxygen demand is elevated by environmental warming.

Gene--environment interactions influence feeding and anti-predator behavior in wild and transgenic coho salmon.

Environmental conditions are known to affect phenotypic development in many organisms, making the characteristics of an animal reared under one set of conditions not always representative of animals reared under a different set of conditions. Previous results show that such plasticity can also affect the phenotypes and ecological interactions of different genotypes, including animals anthropogenically generated by genetic modification. To understand how plastic development can affect behavior in animals of different genotypes, we examined the feeding and risk-taking behavior in growth-enhanced transgenic coho salmon (with two- to threefold enhanced daily growth rates compared to wild type) under a range of conditions. When compared to wild-type siblings, we found clear effects of the rearing environment on feeding and risk-taking in transgenic animals and noted that in some cases, this environmental effect was stronger than the effects of the genetic modification. Generally, transgenic fish, regardless of rearing conditions, behaved similar to wild-type fish reared under natural-like conditions. Instead, the more unusual phenotype was associated with wild-type fish reared under hatchery conditions, which possessed an extreme risk averse phenotype compared to the same strain reared in naturalized conditions. Thus, the relative performance of genotypes from one environment (e.g., laboratory) may not always accurately reflect ecological interactions as would occur in a different environment (e.g., nature). Further, when assessing risks of genetically modified organisms, it is important to understand how the environment affects phenotypic development, which in turn may variably influence consequences to ecosystem components across different conditions found in the complexity of nature.

Accuracy of nonmolecular identification of growth-hormone- transgenic coho salmon after simulated escape.

Concerns with transgenic animals include the potential ecological risks associated with release or escape to the natural environment, and a critical requirement for assessment of ecological effects is the ability to distinguish transgenic animals from wild type. Here, we explore geometric morphometrics (GeoM) and human expertise to distinguish growth-hormone-transgenic coho salmon (Oncorhynchus kisutch) specimens from wild type. First, we simulated an escape of 3-month-old hatchery-reared wild-type and transgenic fish to an artificial stream, and recaptured them at the time of seaward migration at an age of 13 months. Second, we reared fish in the stream from first-feeding fry until an age of 13 months, thereby simulating fish arising from a successful spawn in the wild of an escaped hatchery-reared transgenic fish. All fish were then assessed from 'photographs by visual identification (VID) by local staff and by GeoM based on 13 morphological landmarks. A leave-one-out discriminant analysis of GeoM data had on average 86% (72-100% for individual groups) accuracy in assigning the correct genotypes, whereas the human experts were correct, on average, in only 49% of cases (range of 18-100% for individual fish groups). However, serious errors (i.e., classifying transgenic specimens as wild type) occurred for 7% (GeoM) and 67% (VID) of transgenic fish, and all of these incorrect assignments arose with fish reared in the stream from the first-feeding stage. The results show that we presently lack the skills of visually distinguishing transgenic coho salmon from wild type with a high level of accuracy, but that further development-of GeoM methods could be useful in identifying second-generation,fish from nature as a nonmolecular approach.

Warming alters the body shape of European perch Perca fluviatilis.

The consequences of elevated temperature on body shape were investigated by comparing European perch Perca fluviatilis from the Forsmark area of the Baltic Sea to P. fluviatilis from a nearby Biotest enclosure. The Biotest is a man-made enclosure within the Baltic Sea that has received warm water from a nuclear power plant since 1980, resulting in temperatures that are elevated 5-10 °C relative to the surrounding Baltic Sea. Sampled fish ranged from young-of-the-year to 14 years. Geometric morphometrics and multivariate statistical analysis revealed significant morphological differences between individuals of P. fluviatilis from these two habitats. Most importantly, relative shape changed with size, with small individuals of P. fluviatilis from Biotest being characterized by a deeper body shape and a larger caudal peduncle than the smaller Baltic individuals. In large specimens, smaller differences were found with Biotest individuals being more slender than Baltic individuals. These results show that, in order to have a full understanding of the biological effects of elevated temperatures, studies that cover the entire size range of organisms will be important. Apart from the direct influence of temperature on growth rate and body shape, other ecological factors affected by temperature are discussed as possible contributors to the observed differences between the two populations.

Inbreeding-related trade-offs in stress resistance in the ant Formica exsecta.

Inbred individuals and populations are predicted to suffer from inbreeding depression, especially in times of stress. Under natural conditions, organisms are exposed to more than one stressor at any one time, highlighting the importance of stress resistance traits. We studied how inbreeding- and immunity-related traits are correlated under different dietary conditions in the ant Formica exsecta. Its natural diet varies in the amount and nature of plant secondary compounds and the level of free radicals, all of which require detoxification to maintain organismal homeostasis. We found that inbreeding decreased general antibacterial activity under dietary stress, suggesting inbreeding-related physiological trade-offs.

Risk-taking behaviour may explain high predation mortality of GH-transgenic common carp Cyprinus carpio.

The competitive ability and habitat selection of juvenile all-fish GH-transgenic common carp Cyprinus carpio and their size-matched non-transgenic conspecifics, in the absence and presence of predation risk, under different food distributions, were compared. Unequal-competitor ideal-free-distribution analysis showed that a larger proportion of transgenic C. carpio fed within the system, although they were not overrepresented at a higher-quantity food source. Moreover, the analysis showed that transgenic C. carpio maintained a faster growth rate, and were more willing to risk exposure to a predator when foraging, thereby supporting the hypothesis that predation selects against maximal growth rates by removing individuals that display increased foraging effort. Without compensatory behaviours that could mitigate the effects of predation risk, the escaped or released transgenic C. carpio with high-gain and high-risk performance would grow well but probably suffer high predation mortality in nature.

Growth and endocrine effects of recombinant bovine growth hormone treatment in non-transgenic and growth hormone transgenic coho salmon.

To examine the relative growth, endocrine, and gene expression effects of growth hormone (GH) transgenesis vs. GH protein treatment, wild-type non-transgenic and GH transgenic coho salmon were treated with a sustained-release formulation of recombinant bovine GH (bGH; Posilac). Fish size, specific growth rate (SGR), and condition factor (CF) were monitored for 14 weeks, after which endocrine parameters were measured. Transgenic fish had much higher growth, SGR and CF than non-transgenic fish, and bGH injection significantly increased weight and SGR in non-transgenic but not transgenic fish. Plasma salmon GH concentrations decreased with bGH treatment in non-transgenic but not in transgenic fish where levels were similar to controls. Higher GH mRNA levels were detected in transgenic muscle and liver but no differences were observed in GH receptor (GHR) mRNA levels. In non-transgenic pituitary, GH and GHR mRNA levels per mg pituitary decreased with bGH dose to levels seen in transgenic salmon. Plasma IGF-I was elevated with bGH dose only in non-transgenic fish, while transgenic fish maintained an elevated level of IGF-I with or without bGH treatment. A similar trend was seen for liver IGF-I mRNA levels. Thus, bGH treatment increased fish growth and influenced feedback on endocrine parameters in non-transgenic but not in transgenic fish. A lack of further growth stimulation of GH transgenic fish suggests that these fish are experiencing maximal growth stimulation via GH pathways.

Inside the heads of David and Goliath: environmental effects on brain morphology among wild and growth-enhanced coho salmon Oncorhynchus kisutch.

Transgenic and wild-type individual coho salmon Oncorhynchus kisutch were reared in hatchery and near-natural stream conditions and their brain and structure sizes were determined. Animals reared in the hatchery grew larger and developed larger brains, both absolutely and when controlling for body size. In both environments, transgenics developed relatively smaller brains than wild types. Further, the volume of the optic tectum of both genotypes was larger in the hatchery animals and the cerebellum of transgenics was smaller when reared in near-natural streams. Finally, wild types developed a markedly smaller telencephalon under hatchery conditions. It is concluded that, apart from the environment, genetic factors that modulate somatic growth rate also have a strong influence on brain size and structure.

Are you my mother? Kin recognition in the ant Formica fusca.

In social insects, workers trade personal reproduction for indirect fitness returns from helping their mother rear collateral kin. Colony membership is generally used as a proxy for kin discrimination, but the question remains whether recognition allows workers to discriminate between kin and nonkin regardless of colony affiliation. We investigated whether workers of the ant Formica fusca can identify their mother when fostered with their mother, their sisters, a hetero-colonial queen or hetero-colonial workers. We found that workers always displayed less aggression towards both their mother and their foster queen, as compared to an unfamiliar hetero-colonial queen. In support of this finding, workers maintain their colony hydrocarbon profile regardless of foster regime, yet show modifications when exposed to different environments. This indicates that recognition entails environmental and genetic components, which allow both discrimination of kin in the absence of prior contact and learning of recognition cues based on group membership.

Migration and growth potential of coho salmon smolts: implications for ecological impacts from growth-enhanced fish.

Wild-genotype and growth hormone (GH) transgenic coho salmon (with dramatically enhanced growth potential) were used to examine the influence of genotype, age, body size, growth, and rearing conditions on the onset of seaward migration and to assess the potential consequences of the introduction of such transgenic fish on natural ecosystems and wild populations. When reared from the first feeding stage under naturalized stream conditions, there was no difference in survival or migratory timing between the two genotypes. However, larger fish migrated earlier in the season than smaller fish of both genotypes, and transgenic fish with higher specific spring growth rates migrated earlier in the season than slower-growing transgenic fish. Stream-reared fish of both genotypes also displayed increased migratory activity at dawn and dusk. Fish reared in the hatchery for 3 and 15 months before being released into the stream in August differed in onset of seaward migration due mainly to age (older fish migrated earlier in the season) and genotype (transgenic fish migrated before wild-type in younger fish). Further, hatchery-reared fish showed no diel pattern in activity during migration. In older fish, larger individuals migrated later in the season than small individuals, whereas there was no clear size effect in younger individuals. Thus, although small differences in spring migration timing were observed among groups, seaward migration in coho salmon was largely independent of major shifts in size and growth rate induced by GH transgenesis (i.e., transgenic fish migrated at approximately the normal time in the spring, rather than at the typical size). Further, early rearing conditions had a stronger effect on migratory behavior than did the growth-promoting transgene. Taking into account effects of migratory timing, growth, survival, and differential food conversion efficiencies, these data suggest that transgenic fish escaped from hatcheries would have a greater impact on stream ecosystems during early life compared to escaped wild-type fish. However, this difference may be reduced if rearing occurred in subsequent generations under wild conditions where growth rates of transgenic fish are reduced compared with hatchery conditions.

Effects of temperature and growth hormone on individual growth trajectories of wild-type and transgenic coho salmon Oncorhynchus kisutch.

In this study, individual growth patterns of wild-type and growth-enhanced coho salmon Oncorhynchus kisutch at 8, 12 and 16 degrees C water temperature were followed. Despite large differences among individuals in growth rates, there was generally little variation in the shape of the growth curves among O. kisutch individuals of both genotypes and at all temperatures. Typically, individuals that were relatively large initially were also relatively large at the end of the growth period. The limitation in variation was more pronounced in the growth-enhanced O. kisutch than in the wild type, where the relative size of some individuals reared at 12 and 8 degrees C changed by the end of the trial. As a warmer temperature seems to decrease the plasticity of growth trajectories in wild-type fish, it is possible that global warming will influence the ability of wild fish to adapt their growth to changing conditions.

Fitness and the level of homozygosity in a social insect.

To date very few studies have addressed the effects of inbreeding in social Hymenoptera, perhaps because the costs of inbreeding are generally considered marginal owing to male haploidy whereby recessive deleterious alleles are strongly exposed to selection in males. Here, we present one of the first studies on the effects of queen and worker homozygosity on colony performance. In a wild population of the ant Formica exsecta, the relative investment of single-queen colonies in sexual production decreased with increased worker homozygosity. This may either stem from increased homozygosity decreasing the likelihood of diploid brood to develop into queens or a lower efficiency of more homozygous workers at feeding larvae and thus a lower proportion of the female brood developing into queens. There was also a significant negative association between colony age and the level of queen but not worker homozygosity. This association may stem from inbreeding affecting queen lifespan and/or their fecundity, and thus colony survival. However, there was no association between queen homozygosity and colony size, suggesting that inbreeding affects colony survival as a result of inbred queens having a shorter lifespan rather than a lower fecundity. Finally, there was no significant association between either worker or queen homozygosity and the probability of successful colony founding, colony size and colony productivity, the three other traits studied. Overall, these results indicate that inbreeding depression may have important effects on colony fitness by affecting both the parental (queen) and offspring (worker)generations cohabiting within an ant colony.

Elevated ability to compete for limited food resources by 'all-fish' growth hormone transgenic common carp Cyprinus carpio.

Food consumption, number of movements and feeding hierarchy of juvenile transgenic common carp Cyprinus carpio and their size-matched non-transgenic conspecifics were measured under conditions of limited food supply. Transgenic fish exhibited 73.3% more movements as well as a higher feeding order, and consumed 1.86 times as many food pellets as their non-transgenic counterparts. After the 10 day experiment, transgenic C. carpio had still not realized their higher growth potential, which may be partly explained by the higher frequency of movements of transgenics and the 'sneaky' feeding strategy used by the non-transgenics. The results indicate that these transgenic fish possess an elevated ability to compete for limited food resources, which could be advantageous after an escape into the wild. It may be that other factors in the natural environment (i.e. predation risk and food distribution), however, would offset this advantage. Thus, these results need to be assessed with caution.

Importance of Experimental Environmental Conditions in Estimating Risks and Associated Uncertainty of Transgenic Fish Prior to Entry into Nature.

Salmonids show a high degree of phenotypic plasticity that can differ among genotypes, and this variation is one of the major factors contributing to uncertainty in extrapolating laboratory-based risk assessment data to nature. Many studies have examined the relative growth and survival of transgenic and non-transgenic salmonids, and the results have been highly variable due to genotype × environment interactions. The relative survival of fast- and slow-growing strains can reverse depending on the environment, but it is not clear which specific environmental characteristics are driving these responses. To address this question, two experiments were designed where environmental conditions were varied to investigate the contribution of rearing density, food amount, food type, habitat complexity, and risk of predation on relative growth and survival of fast-growing transgenic and slow-growing wild-type coho salmon. The first experiment altered density (high vs. low) and food amount (high vs. low). Density impacted the relative growth of the genotypes, where transgenic fish grew more than non-transgenic fish in low density streams, regardless of food level. Density also affected survival, with high density causing increased mortality for both genotypes, but the mortality of transgenic relative to non-transgenic fish was lower within the high-density streams, regardless of food level. The second experiment altered habitat complexity (simple vs. complex), food type (artificial vs. natural), amount of food (normal vs. satiation), and risk of predation (present vs. absent). Results from this experiment showed that genotype affected growth and survival, but genotype effects were modulated by one or more environmental factors. The effect of genotype on survival was influenced by all examined environmental factors, such that no predictable trend in relative survival of transgenic versus non-transgenic fry emerged. This was primarily due to variations in survival of non-transgenic fish under different environmental conditions (non-transgenic fry had highest survival in hatchery conditions, and lowest survival in complex conditions with natural food fed at a normal level with or without predators). Transgenic fry survival was only significantly influenced by predator presence. The effects of genotype on mass and length were significantly modulated by food type only. Transgenic fry were able to gain a large size advantage over non-transgenic fish when fed artificial food under all habitat types. These experiments support the observations of dynamic responses in growth and survival depending on the environment, and demonstrate the challenge of applying laboratory-based experiments to risk assessment in nature.

Disruption of seasonality in growth hormone-transgenic coho salmon (Oncorhynchus kisutch) and the role of cholecystokinin in seasonal feeding behavior.

Seasonal variation in daily food intake is a well-documented phenomenon in many organisms including wild-type coho salmon where the appetite is noticeably reduced during periods of decreased day length and low water temperature. This reduction may in part be explained by altered production of cholecystokinin (CCK) and growth hormone (GH). CCK is a hormone produced in the brain and gut that mediates a feeling of satiety and thus has an inhibitory effect on food intake and foraging behaviour. Growth hormone (GH) enhances feeding behaviour and consequently growth, but its production is reduced during winter. The objectives of this study were: first, to compare the seasonal feeding behaviour of wild and GH-transgenic coho salmon; second, to determine the behavioural effect of blocking the action of CCK (by using devazepide) on the seasonal food intake; and third, to measure CCK expression in brain and gut tissues between the two genotypes across seasons. We found that, in contrast to wild salmon, food intake in transgenic salmon was not reduced during winter indicating that seasonal control of appetite regulation has been disrupted by constitutive production of GH in transgenic animals. Blocking of CCK increased food intake in both genotypes in all seasons. The increase was stronger in wild genotypes than transgenic fish; however blocking CCK in wild-type fish in winter did not elevate appetites to levels observed in the summer. The response to devazepide was generally faster in transgenic than in wild salmon with more rapid effects observed during summer than during winter, possibly due to a higher temperature in summer. Overall, a seasonal effect on CCK mRNA levels was observed in telencephalon with levels during winter being higher compared to the summer in wild fish, but with no seasonal effect in transgenic fish. No differences in seasonal CCK expression were found in hypothalamus. Higher levels of CCK were detected in the gut of both genotypes in winter compared to summer. Thus, CCK appears to mediate food intake among seasons in both wild-type and GH-transgenic salmon, and an altered CCK regulation may be responsible at least in part for the seasonal regulation of food intake.

On the Similarity of Pulsating and Accelerating Turbulent Pipe Flows.

The near-wall region of an unsteady turbulent pipe flow has been investigated experimentally using hot-film anemometry and two-component particle image velocimetry. The imposed unsteadiness has been pulsating, i.e., when a non-zero mean turbulent flow is perturbed by sinusoidal oscillations, and near-uniformly accelerating in which the mean flow ramped monotonically between two turbulent states. Previous studies of accelerating flows have shown that the time evolution between the two turbulent states occurs in three stages. The first stage is associated with a minimal response of the Reynolds shear stress and the ensemble-averaged mean flow evolves essentially akin to a laminar flow undergoing the same change in flow rate. During the second stage, the turbulence responds rapidly to the new flow conditions set by the acceleration and the laminar-like behavior rapidly disappears. During the final stage, the flow adapts to the conditions set by the final Reynolds number. In here, it is shown that the time-development of the ensemble-averaged wall shear stress and turbulence during the accelerating phase of a pulsating flow bears marked similarity to the first two stages of time-development exhibited by a near-uniformly accelerating flow. The stage-like time-development is observed even for a very low forcing frequency; ω+=ων/u¯τ2=0.00073 (or equivalently, ls+=2/ω+=52 ), at an amplitude of pulsation of 0.5. Some previous studies have considered the flow to be quasi-steady at ls+=52 ; however, the forcing amplitude has been smaller in those studies. The importance of the forcing amplitude is reinforced by the time-development of the ensemble-averaged turbulence field. For, the near-wall response of the Reynolds stresses showed a dependence on the amplitude of pulsation. Thus, it appears to exist a need to seek alternative similarity parameters, taking the amplitude of pulsation into account, if the response of different flow quantities in a pulsating flow are to be classified correctly.

Targeted delivery of antisense oligonucleotides to pancreatic ß-cells.

Antisense oligonucleotide (ASO) silencing of the expression of disease-associated genes is an attractive novel therapeutic approach, but treatments are limited by the ability to deliver ASOs to cells and tissues. Following systemic administration, ASOs preferentially accumulate in liver and kidney. Among the cell types refractory to ASO uptake is the pancreatic insulin-secreting ß-cell. Here, we show that conjugation of ASOs to a ligand of the glucagon-like peptide-1 receptor (GLP1R) can productively deliver ASO cargo to pancreatic ß-cells both in vitro and in vivo. Ligand-conjugated ASOs silenced target genes in pancreatic islets at doses that did not affect target gene expression in liver or other tissues, indicating enhanced tissue and cell type specificity. This finding has potential to broaden the use of ASO technology, opening up novel therapeutic opportunities, and presents an innovative approach for targeted delivery of ASOs to additional cell types.

Growth and survival trade-offs and outbreeding depression in rainbow trout (Oncorhynchus mykiss).

The purpose of this study was to examine, using a rainbow trout (Oncorhynchus mykiss) model system, the fitness consequences of three generations of introgression of genotypes adapted to two different environments (culture and nature). The experiments also isolated the influence of competitive interactions and risk of predation on the relative growth and survival of the wild and backcrossed lines. Line crosses representing fast-growing pure domestic (D), slow-growing pure wild (W), domestic x wild hybrids (F1), F1 x wild backcrosses (B1), and B1 x wild backcrosses (B2) were generated and reared under (1) culture conditions, (2) seminatural conditions with competition among genotypes, and (3) seminatural conditions under risk of predation. Survival of the fry in a seminatural environment with competition fit an additive model of gene action with the domestic fish having the highest survival and the wild fish the lowest, but under risk of predation outbreeding depression was suggested by low survival of the B2 lines. Evidence of a trade-off in growth and survival under risk of predation along with observations of genetically determined behavioral differences among the strains may provide some explanation for the observed differences in survival among the strains. This information is relevant to improving our evolutionary understanding of the interaction among genomes, and the influence of environment, during hybridization events. Results from this experiment indicate that alteration of phenotype likely played a prominent role in the reduced fitness experienced by progeny produced after three generations of introgression, supporting the theory that disruption of genotypes selected for adaptation to local conditions may be a primary cause of outbreeding depression in species such as salmon.