The effects of Pediococcus acidilactici MA18/5M on growth performance, gut integrity, and immune response using in vitro and in vivo Pacific salmonid models.

Microbial management is central to aquaculture's efficiency. Pediococcus acidilactici MA18/5M has shown promising results promoting growth, modulation of the immune response, and disease resistance in many fishes. However, the mechanisms through which this strain confers health benefits in fish are poorly understood, particularly in Pacific salmonid models. Briefly, the aims of this study were to i) assess the protective effects of P. acidilactici MA18/5M by examining gut barrier function and the expression of tight junction (TJ) and immune genes in vitro and in vivo, and ii) to determine the protective effects of this strain against a common saltwater pathogen, Vibrio anguillarum J382. An in vitro model of the salmonid gut was employed utilizing the cell line RTgutGC. Barrier formation and integrity assessed by TEER measurements in RTgutGC, showed a significant decrease in resistance in cells exposed only to V. anguillarum J382 for 24 h, but pre-treatment with P. acidilactici MA18/5M for 48 h mitigated these effects. While P. acidilactici MA18/5M did not significantly upregulate tight junction and immune molecules, pre-treatment with this strain protected against pathogen-induced insults to the gut barrier. In particular, the expression of ocldn was significantly induced by V. anguillarum J382, suggesting that this molecule might play a role in the host response against this pathogen. To corroborate these observations in live fish, the effects of P. acidilactici MA18/5M was evaluated in Chinook salmon reared in real aquaculture conditions. Supplementation with P. acidilactici MA18/5M had no effect on Chinook salmon growth parameters after 10 weeks. Interestingly, histopathological results did not show alterations associated with P. acidilactici MA18/5M supplementation, indicating that this strain is safe to be used in the industry. Finally, the expression pattern of transcripts encoding TJ and immune genes in all the treatments suggest that variation in expression is more likely to be due to developmental processes rather than P. acidilactici MA18/5M supplementation. Overall, our results showed that P. acidilactici MA18/5M is a safe strain for use in fish production, however, to assess the effects on growth and immune response previously observed in other salmonid species, an assessment in adult fish is needed.

Diploid and triploid Chinook salmon (Oncorhynchus tshawytscha) have altered microRNA responses in immune tissues after infection with Vibrio anguillarum.

Production of sterile fishes through artificial retention of a third set of chromosomes (triploidy) is a sustainable alternative for aquaculture since it reduces escapee pressure on wild populations. However, these fishes have reduced survival in stressful conditions and in response to infection. In this study, the impact of Vibrio anguillarum infection on diploid and triploid Chinook salmon (Oncorhynchus tshawytscha) was investigated to identify if there was any significant immune regulation by microRNAs (miRNA). Small RNAs from hindgut, head kidney, and spleen were sequenced to determine if miRNA transcript abundance was altered due to ploidy and infection in nine-month old full-sibling diploids and triploids. All three tissues had differentially expressed miRNA prior to infection, indicating subtle changes in epigenetic regulation due to increased ploidy. Additionally, miRNA were altered by infection, but there was only a difference in spleen miRNA expression between diploids and triploids at three days of infection. Furthermore, one miRNA (ssa-miR-2188-3p) was confirmed as having an altered response to infection in triploids compared to diploids, implicating potential immune dysregulation due to increased ploidy. The miRNAs identified in this study are predicted to target immune pathways, providing evidence for their importance in regulating responses to pathogens. This study is the first to investigate how increased ploidy alters miRNA expression in response to infection. Additionally, it provides evidence for epigenetic dysregulation in triploid fishes, which may contribute to their poor performance in response to stress.

Hematocrit Is Associated with Thermal Tolerance and Modulated by Developmental Temperature in Juvenile Chinook Salmon.

To evaluate whether oxygen-carrying capacity influences thermal tolerance in fishes, we reared four Chinook salmon families in present-day (+0°C) and possible future (+4°C) temperatures and assessed the response of hematocrit (Hct) to acute temperature stress. In the +4°C treatment, Hct increased above control levels when juvenile fish were exposed to their critical thermal maximum (CTmax). Conversely, no effect of temperature stress on Hct was found in the +0°C treatment. Hct was positively associated with CTmax ([Formula: see text]; [Formula: see text]), contributing to the CTmax of the +4°C treatment being significantly higher than that of the +0°C treatment (mean ± SD, [Formula: see text] and [Formula: see text], respectively). The association between CTmax and Hct found here supports the hypothesis that thermal tolerance is affected by oxygen supply to tissue. Moreover, the developmental plasticity of CTmax and Hct could represent an adaptive mechanism for salmon faced with climate change.

Environmental and genetic determinants of transcriptional plasticity in Chinook salmon.

Variation in gene transcription is widely believed to be the mechanistic basis of phenotypically plastic traits; however, comparatively little is known about the inheritance patterns of transcriptional variation that would allow us to predict its response to selection. In addition, acclimation to different environmental conditions influences acute transcriptional responses to stress and it is unclear if these effects are heritable. To address these gaps in knowledge, we assayed levels of messenger RNA for 14 candidate genes at rest and in response to a 24-h confinement stress for 72 half-sib families of Chinook salmon reared in two different environments (hatchery and semi-natural stream channel). We observed extensive plasticity for mRNA levels of metabolic and stress response genes and demonstrated that mRNA level plasticity due to rearing environment affects mRNA level plasticity in response to stress. These effects have important implications for natural populations experiencing multiple stressors. We identified genotype-by-environment interactions for mRNA levels that were dominated by maternal effects; however, mRNA level response to challenge also exhibited a non-additive genetic basis. Our results indicate that while plasticity for mRNA levels can evolve, predicting the outcome of selection will be difficult. The inconsistency in genetic architecture among treatment groups suggests there is considerable cryptic genetic variation for gene expression.

Prenatal Stress Exposure Generates Higher Early Survival and Smaller Size without Impacting Developmental Rate in a Pacific Salmon.

Prenatal exposure to elevated glucocorticoids can act as a signal of environmental stress, resulting in modifications to offspring phenotype. While "negative" phenotypic effects (i.e., smaller size, slower growth) are often reported, recent research coupling phenotype with other fitness-related traits has suggested positive impacts of prenatal stress. Using captive Chinook salmon (Oncorhynchus tshawytscha), we treated eggs with biologically relevant cortisol levels-low (300 ng mL-1 ), high (1,000 ng mL-1 ), or control (0 ng mL-1 )-to examine the early-life impacts of maternally transferred stress hormones on offspring. Specifically, we measured early survival, rate of development, and multiple measures of morphology. Low and high cortisol dosing of eggs resulted in significantly higher survival compared to controls (37% and 24% higher, respectively). Fish reared from high dose eggs were structurally smaller compared to control fish, but despite this variation in structural size, exposure to elevated cortisol did not impact developmental rate. These results demonstrate that elevations in egg cortisol can positively influence offspring fitness through an increase in early survival while also altering phenotype at a critical life-history stage. Overall, these results suggest that exposure to prenatal stress may not always produce apparently negative impacts on offspring fitness and further proposes that complex phenotypic responses should be examined in relevant environmental conditions.

Indirect genetic effects underlie oxygen-limited thermal tolerance within a coastal population of chinook salmon.

With global temperatures projected to surpass the limits of thermal tolerance for many species, evaluating the heritable variation underlying thermal tolerance is critical for understanding the potential for adaptation to climate change. We examined the evolutionary potential of thermal tolerance within a population of chinook salmon (Oncorhynchus tshawytscha) by conducting a full-factorial breeding design and measuring the thermal performance of cardiac function and the critical thermal maximum (CTmax) of offspring from each family. Additive genetic variation in offspring phenotype was mostly negligible, although these direct genetic effects explained 53% of the variation in resting heart rate (fH). Conversely, maternal effects had a significant influence on resting fH, scope for fH, cardiac arrhythmia temperature and CTmax. These maternal effects were associated with egg size, as indicated by strong relationships between the mean egg diameter of mothers and offspring thermal tolerance. Because egg size can be highly heritable in chinook salmon, our finding indicates that the maternal effects of egg size constitute an indirect genetic effect contributing to thermal tolerance. Such indirect genetic effects could accelerate evolutionary responses to the selection imposed by rising temperatures and could contribute to the population-specific thermal tolerance that has recently been uncovered among Pacific salmon populations.

Additive, non-additive and maternal effects of cytokine transcription in response to immunostimulation with Vibrio vaccine in Chinook salmon (Oncorhynchus tshawytscha).

Estimation of quantitative genetic parameters is important for improving salmonid broodstock management in commercial and government hatcheries. Using a replicated 2 × 2 factorial breeding design (48 families and 192 individuals), we partitioned early immune response transcription variation into additive genetic, non-additive genetic, and maternal components in juvenile Chinook salmon (Oncorhynchus tshawytscha). Transcription of four cytokine genes (IL1, TNF-α, IL-8, IL8-R) and two control genes (IgM and RPS-11) was measured relative to an endogenous control (EF1a) before and 24 h after immune stimulation with Vibrio vaccine. Additive genetic variation was not significant for cytokine transcription and heritability ranged from 0.44 (in pre-challenge IL1) to 0.04 (in post-challenge TNF-α). Non-additive genetic variance was significant in post-challenge IL1 (18 %) and TNF-α (12 %) while maternal effects contributed to pre-challenge cytokine transcription. Cytokine transcription co-expressed within but not between pre- and post-challenge states. The lack of additive genetic effects indicates that cytokine transcription is not a likely candidate for selection programs to improve immune function in Chinook salmon. Our results add to the growing evidence that non-additivity in salmon is common and contributes to our understanding of the genetic architecture of transcription. This indicates that transcription variation may act to maintain genetic variation and facilitate rapid adaptive response in salmonids.

Rapid evolution of egg size in captive salmon.

Captive breeding and release programs, widely used to supplement populations of declining species, minimize juvenile mortality to achieve rapid population growth. However, raising animals in benign environments may promote traits that are adaptive in captivity but maladaptive in nature. In chinook salmon, hatchery rearing relaxes natural selection favoring large eggs, allowing fecundity selection to drive exceptionally rapid evolution of small eggs. Trends toward small eggs are also evident in natural populations heavily supplemented by hatcheries, but not in minimally supplemented populations. Unintentional selection in captivity can lead to rapid changes in critical life-history traits that may reduce the success of supplementation or reintroduction programs.

MATERNAL EFFECTS ON OFFSPRING SIZE: VARIATION THROUGH EARLY DEVELOPMENT OF CHINOOK SALMON.

We performed two breeding experiments with chinook salmon (Oncorhynchus tshawytscha) to explore maternal effects on offspring size. We estimated the magnitude of maternal effects as the differences between sire-offspring and dam-offspring regression slopes. Early in life, offspring size is largely influenced by maternal size, but this influence decreases through early development, with the maternal effect becoming negative at intermediate offspring ages (corresponding to a period of reduced growth of progeny hatching from large eggs) and converging on zero as offspring age. Also, egg size was positively correlated with early survival, but negatively correlated with maternal fecundity.