Climate change can impair bacterial pathogen defences in sablefish via hypoxia-mediated effects on adaptive immunity.

Low-oxygen levels (hypoxia) in aquatic habitats are becoming more common because of global warming and eutrophication. However, the effects on the health/disease status of fishes, the world's largest group of vertebrates, are unclear. Therefore, we assessed how long-term hypoxia affected the immune function of sablefish, an ecologically and economically important North Pacific species, including the response to a formalin-killed Aeromonas salmonicida bacterin. Sablefish were held at normoxia or hypoxia (100% or 40% air saturated seawater, respectively) for 6-16 weeks, while we measured a diverse array of immunological traits. Given that the sablefish is a non-model organism, this involved the development of a species-specific methodological toolbox comprised of qPCR primers for 16 key immune genes, assays for blood antibacterial defences, the assessment of blood immunoglobulin (IgM) levels with ELISA, and flow cytometry and confocal microscopy techniques. We show that innate immune parameters were typically elevated in response to the bacterial antigens, but were not substantially affected by hypoxia. In contrast, hypoxia completely prevented the ∼1.5-fold increase in blood IgM level that was observed under normoxic conditions following bacterin exposure, implying a serious impairment of adaptive immunity. Since the sablefish is naturally hypoxia tolerant, our results demonstrate that climate change-related deoxygenation may be a serious threat to the immune competency of fishes.

Expression of Interleukin-1ß protein in vitro,exvivo and in vivo salmonid models.

Interleukin-1ß (IL-1ß) is one of the first cytokines expressed during immune responses, and its levels are affected by many factors, including stress. To date, it has only been possible to measure IL-1ß transcript (mRNA) expression quantitatively in fish using qPCR. This is because previous studies that measured IL-1ß protein concentrations in these taxa used western blotting, which only provides qualitative data. To advance our knowledge of fish IL-1ß biology, and because post-translational processing plays a critical role in the activation of this molecule, we developed a quantitative enzyme-linked immunosorbent assay (ELISA) to accurately measure the concentration of IL-1ß protein in several cell cultures and in vivo in salmonids. We compared changes in IL-1ß protein levels to the expression of its mRNA. The developed ELISA was quite sensitive and has a detection limit of 12.5 pg/mL. The tools developed, and information generated through this research, will allow for a more accurate and complete understanding of IL-1ß's role in the immune response of salmonids.The assay described here has the potential to significantly advance our ability to assess fish health and immune status.

Atlantic Salmon (Salmo salar) bacterial and viral innate immune responses are not impaired by florfenicol or tetracycline administration.

Antibiotics are used to treat bacterial infections in fish aquaculture, and these drugs can interact with immune cells/the immune system and potentially leave fish vulnerable to viral, fungal, parasitic, or other bacterial infections. However, the effects of antibiotics on fish immunity have largely been overlooked by the aquaculture industry. We tested, at 12 and 20 °C, whether tetracycline and florfenicol (the most commonly used antibiotics in commercial aquaculture), affected the Atlantic salmon's capacity to respond to bacterial or viral stimulation. Atlantic salmon were acclimated to 12 or 20 °C and fed with tetracycline or florfenicol (100 and 10 mg kg of body weight-1 day-1, respectively) medicated feed for 15 or 10 days, respectively. Thereafter, we evaluated their immune function prior to, and after, an intraperitoneal injection of Forte Micro (containing inactivated cultures of Aeromonas salmonicida, Vibrio anguillarum, Vibrio ordalii and Vibrio salmonicida) or the viral mimic polyriboinosinic polyribocytidylic acid (pIC). We measured the transcript expression levels of 8 anti-bacterial and 8 anti-viral putative biomarker genes, and the innate (leukocyte respiratory burst, plasma lysozyme activity and hemolytic activity of the alternative complement pathway) and cellular (relative number of erythrocytes, lymphocytes and thrombocytes, and granulocytes such as monocytes and neutrophils) responses to these challenges. Overall, we only found a few minor effects of either tetracycline or florfenicol on immune gene expression or function at either temperature. Although several studies have reported that antibiotics may negatively affect fish immune responses, our results show that industry-relevant dietary tetracycline and florfenicol treatments do not substantially impact the salmon's innate immune responses. Currently, this is the most comprehensive study on the effects of antibiotics administrated according to industry protocols on immune function in Atlantic salmon.

Influence of Supplemental Dietary Cholesterol on Growth Performance, Indices of Stress, Fillet Pigmentation, and Upper Thermal Tolerance of Female Triploid Atlantic Salmon (Salmo salar).

The salmon aquaculture industry must be proactive at developing mitigation tools/strategies to offset the potential negative impacts of climate change. Therefore, this study examined if additional dietary cholesterol could enhance salmon production at elevated temperatures. We hypothesized that supplemental cholesterol could aid in maintaining cell rigidity, reducing stress and the need to mobilize astaxanthin muscle stores, and improving salmon growth and survival at high rearing temperatures. Accordingly, postsmolt female triploid salmon were exposed to an incremental temperature challenge (+0.2°C day-1) to mimic conditions that they experience in sea cages in the summer, with temperature held at both 16 and 18°C for several weeks [i.e., 3 weeks at 16°C, followed by an increase at 0.2°C day-1 to 18°C (10 days), then 5 weeks at 18°C] to prolong their exposure to elevated temperatures. From 16°C onwards, the fish were fed either a control diet, or one of two nutritionally equivalent experimental diets containing supplemental cholesterol [+1.30%, experimental diet #1 (ED1); or +1.76%, experimental diet #2 (ED2)]. Adding cholesterol to the diet did not affect the salmon's incremental thermal maximum (ITMax), growth, plasma cortisol, or liver stress-related transcript expression. However, ED2 appeared to have a small negative impact on survival, and both ED1 and ED2 reduced fillet "bleaching" above 18°C as measured using SalmoFan™ scores. Although the current results suggest that supplementing salmon diets with cholesterol would have few/minimal benefits for the industry, ≤ 5% of the female triploid Atlantic salmon used in this study irrespective of diet died before temperature reached 22°C. These latter data suggest that it is possible to produce all female populations of reproductively sterile salmon that can withstand summer temperatures in Atlantic Canada.

Phenotypic stress response does not influence the upper thermal tolerance of male Atlantic salmon (Salmo salar).

Fish can be identified as either low responders (LR) or high responders (HR) based on post-stress cortisol levels and whether they exhibit a proactive or reactive stress coping style, respectively. In this study, male Atlantic salmon (Salmo salar) from 17 families reared at 9 °C were repeatedly exposed to an acute handling stress over a period of four months, with plasma cortisol levels measured at 1 h post-stress. Fish were identified as either LR or HR if the total Z-score calculated from their cortisol responses fell into the lower or upper quartile ranges, respectively; with intermediate responders (IR) classified as the remainder. Salmon characterized as LR, IR or HR were then subjected to an incremental thermal challenge, where temperature was raised at 0.2 °C day-1 from their acclimation temperature (12 °C) to mimic natural sea-cage farming conditions during the summer in Newfoundland. Interestingly, feed intake remained high up to 22 °C, while previous studies have shown a decrease in salmon appetite after ∼16-18 °C. After the first three mortalities were recorded at elevated temperature, a subset of LR and HR salmon were exposed to another acute handling stress event at 23.6 °C. Basal and post-stress measurements of plasma cortisol, glucose and lactate did not differ between stress response phenotypes at this temperature. In the end, the average incremental thermal maximum (ITMax) of LR and HR fish was not different (25.1 °C). In comparison, the critical thermal maximum (CTMax; temperature increased at 2 °C h-1) of the remaining IR fish that had been held at 12 °C was 28.5 °C. Collectively, these results: 1) show that this population of Atlantic salmon is very thermally tolerant, and further question the relevance of CTMax in assessing responses to real-world temperature changes; and 2) indicate that characterization of stress phenotype at 9 °C is not predictive of their stress response or survival at high temperatures. Therefore, selection of fish based on phenotypic stress response at low temperatures may not be beneficial to incorporate into Atlantic salmon breeding programs, especially if the goal is to improve growth performance and survival at high temperatures in sea-cages.

The Atlantic salmon's stress- and immune-related transcriptional responses to moderate hypoxia, an incremental temperature increase, and these challenges combined.

The marine environment is predicted to become warmer, and more hypoxic, and these conditions may negatively impact the health and survival of coastal fish species, including wild and farmed Atlantic salmon (Salmo salar). Thus, we examined how: (1) moderate hypoxia (∼70% air saturation) at 12°C for 3 weeks; (2) an incremental temperature increase from 12°C to 20°C (at 1°C week-1) followed by 4 weeks at 20°C; and (3) treatment "2" combined with moderate hypoxia affected transcript expression in the liver of post-smolts as compared to control conditions (normoxia, 12°C). Specifically, we assessed the expression of 45 genes related to the heat shock response, oxidative stress, apoptosis, metabolism and immunity using a high-throughput qPCR approach (Fluidigm Biomark™ HD). The expression profiles of 27 "stress"-related genes indicated that: (i) moderate hypoxia affected the expression of several stress genes at 12°C; (ii) their expression was impacted by 16°C under normoxic conditions, and this effect increased until 20°C; (iii) the effects of moderate hypoxia were not additive to those at temperatures above 16°C; and (iv) long-term (4 weeks) exposure to 20°C, with or without hypoxia, resulted in a limited acclimatory response. In contrast, the expression of 15 immune-related genes was not greatly affected until temperatures reached 20°C, and this effect was particularly evident in fish exposed to the added challenge of hypoxia. These results provide valuable information on how these two important environmental factors affect the "stress" physiology and immunology of Atlantic salmon, and we identify genes that may be useful as hypoxia and/or temperature biomarkers in salmonids and other fishes.

The transcriptomic responses of Atlantic salmon (Salmo salar) to high temperature stress alone, and in combination with moderate hypoxia.

BACKGROUND: Increases in ocean temperatures and in the frequency and severity of hypoxic events are expected with climate change, and may become a challenge for cultured Atlantic salmon and negatively affect their growth, immunology and welfare. Thus, we examined how an incremental temperature increase alone (Warm & Normoxic-WN: 12 → 20 °C; 1 °C week- 1), and in combination with moderate hypoxia (Warm & Hypoxic-WH: ~ 70% air saturation), impacted the salmon's hepatic transcriptome expr\ession compared to control fish (CT: 12 °C, normoxic) using 44 K microarrays and qPCR. RESULTS: Overall, we identified 2894 differentially expressed probes (DEPs, FDR < 5%), that included 1111 shared DEPs, while 789 and 994 DEPs were specific to WN and WH fish, respectively. Pathway analysis indicated that the cellular mechanisms affected by the two experimental conditions were quite similar, with up-regulated genes functionally associated with the heat shock response, ER-stress, apoptosis and immune defence, while genes connected with general metabolic processes, proteolysis and oxidation-reduction were largely suppressed. The qPCR assessment of 41 microarray-identified genes validated that the heat shock response (hsp90aa1, serpinh1), apoptosis (casp8, jund, jak2) and immune responses (apod, c1ql2, epx) were up-regulated in WN and WH fish, while oxidative stress and hypoxia sensitive genes were down-regulated (cirbp, cyp1a1, egln2, gstt1, hif1α, prdx6, rraga, ucp2). However, the additional challenge of hypoxia resulted in more pronounced effects on heat shock and immune-related processes, including a stronger influence on the expression of 14 immune-related genes. Finally, robust correlations between the transcription of 19 genes and several phenotypic traits in WH fish suggest that changes in gene expression were related to impaired physiological and growth performance. CONCLUSION: Increasing temperature to 20 °C alone, and in combination with hypoxia, resulted in the differential expression of genes involved in similar pathways in Atlantic salmon. However, the expression responses of heat shock and immune-relevant genes in fish exposed to 20 °C and hypoxia were more affected, and strongly related to phenotypic characteristics (e.g., growth). This study provides valuable information on how these two environmental challenges affect the expression of stress-, metabolic- and immune-related genes and pathways, and identifies potential biomarker genes for improving our understanding of fish health and welfare.

Research on sablefish (Anoplopoma fimbria) suggests that limited capacity to increase heart function leaves hypoxic fish susceptible to heat waves.

Studies of heart function and metabolism have been used to predict the impact of global warming on fish survival and distribution, and their susceptibility to acute and chronic temperature increases. Yet, despite the fact that hypoxia and high temperatures often co-occur, only one study has examined the effects of hypoxia on fish thermal tolerance, and the consequences of hypoxia for fish cardiac responses to acute warming have not been investigated. We report that sablefish (Anoplopoma fimbria) did not increase heart rate or cardiac output when warmed while hypoxic, and that this response was associated with reductions in maximum O2 consumption and thermal tolerance (CTmax) of 66% and approximately 3°C, respectively. Further, acclimation to hypoxia for four to six months did not substantially alter the sablefish's temperature-dependent physiological responses or improve its CTmax. These results provide novel, and compelling, evidence that hypoxia can impair the cardiac and metabolic response to increased temperatures in fish, and suggest that some coastal species may be more vulnerable to climate change-related heat waves than previously thought. Further, they support research showing that cross-tolerance and physiological plasticity in fish following hypoxia acclimation are limited.

Dietary ß-glucan (MacroGard®) improves innate immune responses and disease resistance in Nile tilapia regardless of the administration period.

The effects of dietary ß-glucan on innate immune responses have been shown in a number of different vertebrate species. However, there is conflicting information about the period of administration (shorter vs. longer), and it is also unclear to what extent ß-glucan's effects can be observed post-treatment in fish. Thus, we fed Nile tilapia for 0 (control group; 45 days of control diet), 15 (30 days of control followed by 15 days of ß-glucan), 30 (15 days of control followed by 30 days of ß-glucan) or 45 days with a diet containing 0.1% of ß-glucan (MacroGard®). We evaluated the growth performance at the end of the ß-glucan feeding trial and the innate immune function immediately after the feeding trial and 7 and 14 days post-feeding trial. In addition, at day 10 post-feeding trial, we assessed the tilapia's resistance against a bacterial infection. No significant differences were observed in growth performance between the groups; however, fish fed with ß-glucan for 30 and 45 days had higher (approx. 8%) relative weight gain compared to the control. Regardless of the administration period, fish fed with ß-glucan had higher innate immune responses immediately after the feeding trial such as lysozyme activity in plasma, liver and intestine and respiratory burst compared to the control, and in general these differences were gradually reduced over the withdrawal period (up to 14 days). No differences were observed in the plasma hemolytic activity of the complement or myeloperoxidase activity in plasma or intestine. Moreover, fish from the control group had early mortalities (2 vs. 4-5 days post-infection, respectively) and a lower survival rate (60 vs. 80%, respectively) compared to fish fed with ß-glucan for 15 or 30 days, and, interestingly, fish fed for 45 days with ß-glucan had no mortality. This study indicates that regardless of the administration period (i.e., 15 up to 45 days), the ß-glucan improved the innate immune responses and the tilapia's resistance to disease, and this protection could be observed up to 10 days post-feeding trial, adding in vivo evidence that ß-glucan may contribute to a trained innate immunity. Additionally, we showed that a longer period of administration did not cause immunosuppression as previously hypothesized but promoted further growth and immune performance. These findings are relevant to the aquaculture industry and demonstrate that a longer ß-glucan feeding protocol may be considered to achieve better results.

Development of Fish Immunity and the Role of ß-Glucan in Immune Responses.

Administration of ß-glucans through various routes, including immersion, dietary inclusion, or injection, have been found to stimulate various facets of immune responses, such as resistance to infections and resistance to environmental stress. ß-Glucans used as an immunomodulatory food supplement have been found beneficial in eliciting immunity in commercial aquaculture. Despite extensive research involving more than 3000 published studies, knowledge of the receptors involved in recognition of ß-glucans, their downstream signaling, and overall mechanisms of action is still lacking. The aim of this review is to summarize and discuss what is currently known about of the use of ß-glucans in fish.

The Innate Immune Response of Atlantic Salmon (Salmo salar) Is Not Negatively Affected by High Temperature and Moderate Hypoxia.

Climate change is predicted to increase water temperatures and decrease oxygen levels in freshwater and marine environments, however, there is conflicting information regarding the extent to which these conditions may impact the immune defenses of fish. In this study, Atlantic salmon were exposed to: (1) normoxia (100-110% air saturation) at 12°C; (2) an incremental temperature increase (1°C per week from 12 to 20°C), and then held at 20°C for an additional 4 weeks; and (3) "2" with the addition of moderate hypoxia (~65-75% air saturation). These conditions realistically reflect what farmed salmon in some locations are currently facing, and future conditions in Atlantic Canada and Europe, during the summer months. The salmon were sampled for the measurement of head kidney constitutive anti-bacterial and anti-viral transcript expression levels, and blood parameters of humoral immune function. Thereafter, they were injected with either the multi-valent vaccine Forte V II (contains both bacterial and viral antigens) or PBS (phosphate-buffer-saline), and the head kidney and blood of these fish were sampled at 6, 12, 24, and 48 h post-injection (HPI). Our results showed that: (1) neither high temperature, nor high temperature + moderate hypoxia, adversely affected respiratory burst, complement activity or lysozyme concentration; (2) the constitutive transcript expression levels of the anti-bacterial genes il1ß, il8-a, cox2, hamp-a, stlr5-a, and irf7-b were up-regulated by high temperature; (3) while high temperature hastened the peak in transcript expression levels of most anti-bacterial genes by 6-12 h following V II injection, it did not affect the magnitude of changes in transcript expression; (4) anti-viral (viperin-b, mx-b, and isg15-a) transcript expression levels were either unaffected, or downregulated, by acclimation temperature or V II injection over the 48 HPI; and (5) hypoxia, in addition to high temperature, did not impact immune transcript expression. In conclusion, temperatures up to 20°C, and moderate hypoxia, do not impair the capacity of the Atlantic salmon's innate immune system to respond to bacterial antigens. These findings are surprising, and highlight the salmon's capacity to mount robust innate immune responses (i.e., similar to control fish under optimal conditions) under conditions approaching their upper thermal limit.

ß-Glucan enhances respiratory activity of leukocytes suppressed by stress and modulates blood glucose levels in pacu (Piaractus mesopotamicus).

We evaluated the immune response of pacu fed with a ß-glucan diet (0.5%) for 10 days. After the feeding period, fish were subjected to handling and 3 h after, inoculated with Aeromonas hydrophila. Fish were sampled before handling (baseline condition), 3, 6, and 24 h and 1 week after inoculation. A higher level of blood glucose was found in fish treated with ß-glucan in baseline conditions. Handling and bacterial inoculation increased the circulating levels of cortisol and glucose and promoted the acute inflammatory response (lymphopenia and neutrophilia). ß-Glucan prevented the decrease in the respiratory activity of leukocytes observed in the control group at 3 h sampling. ß-Glucan did not affect the complement and lysozyme, which were activated 24 h after the bacterial challenge in control fish. A reduction in the number of leukocytes was found in fish treated with ß-glucan 1 week after the challenge. We suggest two plausible hypotheses for this event: (1) it could be attributed to a depletion of the immune responses or (2) it could be due to a mobilization of the leukocytes to the spleen for antigen presenting/processing. In general, ß-glucan avoided the reduction of the activity of leukocytes after stress and the bacterial challenge and increased the baseline glucose levels. Our findings confirm the immunomodulatory action of glucan and add evidence showing that glucan can have a role in stress response.

Intermediary metabolic response and gene transcription modulation on the Sub-Antarctic notothenioid Eleginops maclovinus (Valenciennes, 1930) injected with two strains of Piscirickettsia salmonis.

Pathogen interactions with cultured fish populations are well studied, but their effects on native fishes have not been characterized. In Chile, the disease caused by bacterial species Piscirickettsia salmonis represents one of the main issues and is considered to be one of the important pathogens in the field of aquaculture. They have been found to infect native fish. Therefore, it is necessary to understand the impact of P. salmonis on native species of local commercial value, as well as the potential impact associated with the emergence of antibiotic-resistant strains of P. salmonis. Due to this purpose, the native fish Eleginops maclovinus was used in our study. Fish were randomly distributed in tanks and intraperitoneally inoculated with two strains of P. salmonis. No mortality was recorded during the experiment. Cortisol, glucose and total α-amino acid levels increased in fish injected with AUSTRAL-005 strain compared to sham-injected and LF-89-inoculated fish. Moreover, results showed an increase in the activity of carbohydrates and lipids metabolism in liver; and an increase in the carbohydrates, lipids and total α-amino acid metabolism in muscle after injection with AUSTRAL-005. Our results suggest that P. salmonis modulates the physiology of E. maclovinus and the physiological impact increase in the presence of the antibiotic-resistant strain AUSTRAL-005.

ß-glucan modulates cortisol levels in stressed pacu (Piaractus mesopotamicus) inoculated with heat-killed Aeromonas hydrophila.

In this study, we show that ß-glucan can modulate cortisol release in fish. We simulated a common situation in aquaculture: the transport of fish followed by contact with an opportunistic pathogen and observed what effect glucan had on the immune and stress response in these conditions. Pacu (Piaractus mesopotamicus) were fed with a diet containing ß-glucan (0.1%) for 15 days prior to transport followed by an injection with heat-killed Aeromonas hydrophila. We sampled fish before transport, at arrival and at 3 and 24 h after bacterial injection. ß-Glucans are used in aquaculture and have a known immunostimulatory effect, which was observed in this study. The results showed that ß-glucan modulated the plasma cortisol levels differently by increasing these levels up to 24 h after transport and preventing the increase caused by bacterial inoculum injection. In addition, ß-glucan enhanced the activity of the complement system at 24 h and reduced the monocytes and lymphocytes number in peripheral blood at 3 and 24 h after bacterial inoculation. Our results suggest that ß-glucan modulated a bidirectional interaction between the stress and the immune responses. The modulation of cortisol levels and the immunostimulation by ß-glucan at different moments in our study suggest the compound has a protective effect by avoiding higher levels of the hormone and improving resistance against bacterial infection in pacu. These results add evidence to support the use of ß-glucan as an immunomodulator in the aquaculture industry.

Innate response based on visual cues of sympatric and allopatric predators in Nile tilapia.

Predators are an important selective pressure for prey, and responses to visual exposure to non-predators and to allopatric and sympatric predators may assist the understanding of how prey animals recognize and distinguish potential threats. Here, we visually exposed predator-naïve Nile tilapia (Oreochromis niloticus) to aquaria with only water (handling-negative control), a non-predator fish (Cyprinus carpio), a South American catfish (Pseudoplatystoma coruscans, allopatric predator), and an African catfish (Clarias gariepinus; sympatric predator). We found that Nile tilapia displayed antipredator responses to both predator catfish but not to non-predator fish or negative control conditions. These antipredator responses, however, were stronger to the sympatric catfish in comparison to the allopatric animal. We concluded that Nile tilapia is able to innately distinguish between a sympatric and an allopatric predator based on visual cues. We assume that the innate recognition of the allopatric predator could be attributed to similarities in the body morphology and movement (key stimuli) of the South American and African catfish. Although this is plausible, the mechanism of the innate allopatric recognition remains unknown and deserves future investigation.

The acute and incremental thermal tolerance of Atlantic cod (Gadus morhua) families under normoxia and mild hypoxia.

Given climate change projections, the limited ability of fish reared in sea-cages to behaviourally thermoregulate, and that thermal tolerance may be heritable, studies that examine family-related differences in upper thermal tolerance are quite relevant to the aquaculture industry. Thus, we investigated the upper thermal tolerance of 15 Atlantic cod (Gadus morhua L.) families by challenging them with acute (2 °C h-1) and incremental (1 °C every 4 days) temperature increases (CTmax and ITmax tests, respectively) under normoxia (~ 100% air saturation) and mild hypoxia (~ 75% air sat.). The cod's CTmax was 22.5 ±â€¯0.1 °C (mean ±â€¯S.E.) during normoxia and 21.8 ±â€¯0.1 °C during hypoxia (P < 0.001); and these two CTmax values were significantly correlated across families. In both the normoxic and hypoxic ITmax tests, feed intake fell by ~50% between 17 and 18 °C, and stopped entirely by 21 °C. No mortalities were observed under 20 °C in the normoxic and hypoxic ITmax tests, and the ITmax value was ~21.7 °C in both groups. Differences in the upper thermal tolerance between families were only observed in the CTmax experiment. No correlation was found between the specific growth rate and the CTmax of the families. Further, no correlation existed between CTmax and ITmax. This study is the first to compare the thermal tolerance of fish families to both CTmax and ITmax challenges, and the data: 1) suggest that the Atlantic cod is quite tolerant of acute (i.e., hours) or short-term (i.e., weeks) exposure to high water temperatures (i.e., up to 20 °C); 2) indicate that it might be difficult to select fish with higher ITmax values; and 3) question the relevance of CTmax for selecting fish that are destined for sea-cages where temperatures slowly warm over the summer.

Modulation of stress and innate immune response by corticosteroids in pacu (Piaractus mesopotamicus).

Understanding how stress and corticosteroid modulates the innate immune response is one of the keys to improving productivity and reducing losses in intensive aquaculture. Thus, we investigated the effects of dietary corticosteroids (7 days; long-term exposure) and transport (4 h; short-term stress) on stress and innate immune response in pacu. For this end, fish were fed with diets containing dexamethasone (100 mg kg-1) or hydrocortisone (200 mg kg-1), followed by transport, and then were intraperitoneally inoculated with heat-killed Aeromonas hydrophila or PBS (sham-inoculation). Fish were sampled after a 7-day feeding period, immediately post-transport and 24 h post-transport and inoculation. The dietary treatment of corticosteroids decreased resting cortisol levels by inhibiting the production of cortisol on the hypothalamus pituitary interrenal-axis. Further, both corticosteroids reduced hematocrit, red blood cells, haemoglobin and hemolytic activity of the complement, while they increased glucose levels and serum lysozyme concentrations. The transport increased cortisol and glucose levels and reduced the humoral immune defenses such as serum lysozyme concentration and hemolytic activity of the complement system. Interestingly, the hemolytic activity of the complement system increased sharply in fish fed with corticosteroids immediately post-transport, when they had their HPI-axis partially suppressed by the corticosteroids. This finding suggests a stimulatory effect of the catecholamines released during the transport on the activity of the complement system. Our results are highly valuable to understanding the stress and innate immune responses to long-term exposure to corticosteroids and short-term stress in fish and may provide insights into how corticosteroids modulate the innate immune system.

The environmental tolerances and metabolic physiology of sablefish (Anoplopoma fimbria).

Given the potential impacts of global warming, such as increases in temperature and the frequency/severity of hypoxia in marine ecosystems, it is important to study the impacts of these environmental challenges on sea-cage reared aquaculture species. This study focuses on the sablefish (Anoplopoma fimbria), an emerging aquaculture species that has a unique ecology in the wild. For instance, adults inhabit oxygen minimum zones and cool waters at depths up to 1500 m. Using Atlantic salmon (Salmo salar) (~1132 g adults) as a comparative species, we used intermittent-flow respirometry to characterize the tolerance and metabolic response of sablefish (~10 g juveniles and ~675 g adults) to acute increases in temperature (2 °C h-1) and decreases in oxygen level (~10% air saturation h-1). Adult sablefish were much more hypoxia tolerant than adult salmon [O2 level at loss of equilibrium ~5.4% vs. ~24.2% air saturation, respectively]. In addition, sablefish could withstand upper temperatures only slightly lower than salmon [critical thermal maximum (CTmax) ~24.9 °C vs. ~26.2 °C, respectively]. Sablefish juveniles were both less hypoxia and thermally tolerant than adults [critical O2 tension ~18.9% vs. ~15.8% air saturation; CTmax ~22.7 vs. ~24.9 °C, respectively]. Interestingly, many of these differences in environmental tolerance could not be explained by differences in metabolic parameters (aerobic scope or routine metabolic rate). Our findings show that sablefish are tolerant of high temperatures, and very tolerant of hypoxia, traits that are advantageous for an aquaculture species in the era of climate change.

ß-Glucan successfully stimulated the immune system in different jawed vertebrate species.

Several reports have shown the positive effects of ß-glucans on the immune. Howeverthese studies have a broad experimental design including ß-glucans compounds. Consequently, a study using the same ß-glucan molecule, administration route and experimental design is needed to compare the effects of ß-glucan across vertebrate species. For this end, during 28 days we fed four different vertebrate species: mice, dogs, piglets and chicks, with two ß-glucan molecules (BG01 and BG02). We measured the serum interleukin 2 as an indicator of innate immune response, the neutrophils and monocytes phagocytosis index as a cellular response and antibody formation as an adaptive response. The results clearly showed that the different ß-glucan molecules exhibited biologically differently behaviors, but both molecules stimulate the immune system in a similar pattern in these four species. This finding suggests that vertebrates shared similar mechanisms/patterns in recognizing the ß-glucans and confirms the benefits of ß-glucans across different vertebrate species.

Immunomodulation of Juvenile Pacu, Piaractus mesopotamicus, by Different ß(1-3)(1-6)-D glucan Products

Abstract Stress in intensive fish farming hamper immune function of fish and cause losses by disease outbreaks, a situation that can be minimized, but cannot be completely circumvented, by the use of immunomodulators. Addition of immunomodulators to aquafeeds has thus become a common practice. β-glucan (BG) is one of most studied and effective immunomodulators, aquaculture purposes included. Extracted from cell walls of bacteria, fungi and selected cereals, BG activity depends on the source and extraction methods. This study evaluated effects of two BG products (BG1 and BG2), extracted from Saccharomyces cerevisiae under varying extraction methods and with different immune activity, on the feeding of pacu Piaractus mesopotamicus juveniles. BG1 provided higher leukocytes respiratory activity when fed at 0.5% inclusion for 10 days and 0.1% inclusion for 15 days. Both products seems to cause negative effect on lysozyme concentration and monocytes profile when fed to pacu for 15 days at 0.5% inclusion. Although the results for BG2 did not differ from control (diet devoid of BG), the proximity with the BG1 behavior is a indicative that a commercial product with smaller BG concentration can be effective when more refined technology is used in extraction process.