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Fish in the wild often contend with complex flows that are produced by natural and artificial structures. Research into fish interactions with turbulence often investigates metrics such as turbulent kinetic energy (TKE) or fish positional location, with less focus on the specific interactions between vortex organization and body swimming kinematics. Here, we compared the swimming kinematics of rainbow trout (Oncorhynchus mykiss) holding station in flows produced by two different 3×5 cylinder arrays. We systematically utilized computational fluid dynamics to identify one array that produced a Kármán vortex street with high vortex periodicity (KVS array) and another that produced low periodicity, similar to a parallel vortex street (PVS array), both validated with particle image velocimetry. The only difference in swimming kinematics between cylinder arrays was an increased tail beat amplitude in the KVS array. In both cylinder arrays, the tail beat frequency decreased and snout amplitude increased compared with the freestream. The center of mass amplitude was greater in the PVS array than in only the freestream, however, suggesting some buffeting of the body by the fluid. Notably, we did not observe Kármán gaiting in the KVS array as in previous studies. We hypothesize that this is because (1) vorticity was dissipated in the region where fish held station or (2) vortices were in-line rather than staggered. These results are the first to quantify the kinematics and behavior of fishes swimming in the wake of multiple cylinder arrays, which has important implications for biomechanics, fluid dynamics and fisheries management., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2024. Published by The Company of Biologists Ltd.)

Habitat selection theory enables inferences about species habitat choice across a range of observed population densities. However, it is relatively uncommon to use habitat selection theory in studies of habitat restoration efficacy to understand the effect of restoration on habitat competition. We combined observational density data and resource selection functions to analyze habitat correlations with both habitat selection theory and a mark-recapture experiment to show how habitat restoration can mitigate competition between species with similar habitat preferences. To restore degraded and channelized riverine habitat for juvenile Chinook salmon (Oncorhynchus tshawytscha) and steelhead trout (Oncorhynchus mykiss) engineered log jams (ELJs) have been installed to create pools to enhance growth and rearing. Application of habitat selection theory first showed that both species share a preference for ELJ-treated habitat over unrestored habitat. Linear models showed that steelhead are generalists with respect to depth in unrestored habitat, whereas both species' abundance varies along a depth gradient in ELJ-treated habitat. Selective versus opportunistic use of deep and shallow ELJ pools was density-dependent. We found a range of densities at which a "ghost of competition" exists, where Chinook are selective on deep ELJ-treated pools and steelhead are selective on shallow pools. A mark-recapture experiment confirmed that steelhead limit Chinook movement into unrestored habitat, but this competitive effect vanished in ELJ-treated habitat where selection occurred with respect to pool depth. The experiment, combined with theory, enabled (1) the identification of a mechanism allowing for shared preference of restored habitat and (2) the description of how restoration can mitigate competition., (© 2024 The Ecological Society of America.)

No current technology can specifically target grass carp (Ctenopharyngodon idella) for control within aquatic ecosystems. Rotenone and Carbon Dioxide-Carp are currently the only available registered pesticides for grass carp; they are nonselective and typically applied throughout the water, equally exposing target and native species. A more selective control tool or pesticide application could be used by resource managers to support mitigation efforts. Development of delivery systems that exploit carp feeding strategies could increase selectivity of pesticides and minimize effects on native fishes. A pesticide with selective delivery could be less labor intensive and used within an integrative pest management strategy. The present study examined Antimycin A toxicity in juvenile and sub-adult grass carp and rainbow trout (Oncorhynchus mykiss) across two routes of exposure. Water-based toxicity studies were used to calculate the concentration to cause lethality in 50% of treated fish (LC 50 ) at 24-h, while oral gavage toxicity studies were used to calculate the dose to cause lethality in 50% of treated grass carp and rainbow trout (LD 50 ) 24- to 96-h. Although rainbow trout were more sensitive than grass carp to Antimycin A through water-based exposure, oral toxicity was similar between species, even with inherent gastrointestinal morphological differences. Successful delivery of a lethal dose of Antimycin A to grass carp was achieved through an oral route of exposure using the rapeseed bait and shows promise for registration as a control tool and eventual use in pest management plans. Although a lethal dose of Antimycin A could be incorporated into a single bait pellet, more bait was required to achieve desired mortality when fed to fish under laboratory conditions., (© 2024. Brianne M. Korducki. Parts of this work were authored by US Federal Government authors and are not under copyright protection in the US; foreign copyright protection may apply.)

Increasing evidence shows that larger fish are more vulnerable to acute warming than smaller individuals of the same species. This size-dependency of thermal tolerance has been ascribed to differences in aerobic performance, largely owing to a decline in oxygen supply relative to demand. To shed light on these ideas, we examined metabolic allometry in 130 rainbow trout ranging from 12 to 358 g under control conditions (17°C) and in response to acute heating (to 25°C), with and without supplemental oxygen (100% versus 150% air saturation). Under normoxia, high temperature caused an average 17% reduction in aerobic scope compared with 17°C. Aerobic performance disproportionally deteriorated in bigger fish as the scaling exponent (b) for aerobic scope declined from b=0.87 at 17°C to b=0.74 at 25°C. Hyperoxia increased maximum metabolic rate and aerobic scope at both temperatures and disproportionally benefited larger fish at 25°C as the scaling exponent for aerobic scope was reestablished to the same level as at 17°C (b=0.86). This suggests that hyperoxia may provide metabolic refuge for larger individuals, allowing them to sustain aerobic activities when facing acute warming. Notably, the elevated aerobic capacity afforded by hyperoxia did not appear to improve thermal resilience, as mortality in 25°C hyperoxia (13.8%, n=4) was similar to that in normoxia (12.1%, n=4), although we caution that this topic warrants more targeted research. We highlight the need for mechanistic investigations of the oxygen transport system to determine the consequences of differential metabolic scaling across temperature in a climate warming context., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2024. Published by The Company of Biologists Ltd.)

The negative effects of global warming also directly affect aquatic populations. Consequences such as evaporation due to chronic temperature increase, increase in salinity, and increase in stock density per unit volume are potential stress factors. While creating the trial design, an attempt was made to simulate the effects of global warming, especially on species living in salty and brackish water biotopes. In this study, changes in the gills of rainbow trout (Oncorhynchus mykiss) acclimated to 0, 20, and 38 ‰ of saline in the laboratory were examined histologically and immunohistochemically and blood serum osmolarity. In addition, the water temperature was changed, and experiments were carried out at 16, 19, and 22 °C for each salinity group in parallel with the increase in salinity. However, to simulate the decrease in water volume and intensive stocking due to the potential impact of climate change, the study was carried out using 15 fishes in low-volume aquariums (45 L). Tap water that had been kept for at least 3 days was used in the aquariums. To protect the water quality, independent aquariums with sponge filters were used, and since the aim was to keep dissolved oxygen low, no ventilation system other than the sponge filter was used. In order to minimize the deterioration in water quality during the trial, a 15% water change was performed by performing a bottom flush every 4 days and water of the same temperature and salinity was added as much as the reduced volume. In addition, since increasing stock density due to temperature increase and water decrease will cause the amount of dissolved oxygen to decrease, pure oxygen was not entered into any tank throughout the experiment, and the concentration was requested to be at a low level (7 ± 0.13 mg/L) in all groups. The trials were terminated at the end of the 71st day. Increased serum osmolarity values were observed due to the increase in salinity, and the highest serum osmolarity value was measured at 644 mOsm/kg in the 38 ‰ salinity group. Differences between the groups were found to be statistically significant (p < 0.05). It was observed that the number of cells containing Na + /K + -ATPase increased depending on salinity. Also, the number of chloride cells reached the maximum level in the 38 ‰ salinity group. Due to increasing salt levels, an increase in mucus cells, limited onset hyperplasia, aneurysm, lamellar separation, and necrosis were observed in the gill tissue., Competing Interests: Declarations. Ethics approval: All procedures performed on fish were carried out in accordance with the protocols approved by Van Yüzüncü Yıl University Animal Research Local Ethics Committee (protocol no. 2023/08–06). Consent to participate: Not applicable. Consent for publication: Not applicable. Conflict of interest: The authors declare no competing interests., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

In the present study, we aimed to determine the parameters of oxidative stress markers, motility and kinematics of rainbow trout (Oncorhynchus mykiss) sperm cells exposed to different doses (0.001, 0.01, 0.1, 1.0, and 10 mg/L, in vitro 4 h) of tire rubber based microplastic particles (TRMP-Ps) the leachates procedure of rubber pieces. First of all, TRMP-Ps were prepared by abrasion method in accordance with the literature. Structural and morphological features of TRMP-Ps were determined by Fourier transform infrared (FTIR) spectroscopy and scanning electron microscopy (SEM) methods, respectively. Energy dispersive X-ray (EDX) analysis technique was used to characterize the elemental composition of TRMP-Ps. Particle size of microplastic structures was measured hydrodynamically with dynamic light scattering analysis (DLS). After exposure, the effect of TRMP-Ps was defined by the observations of kinematics and antioxidant activities in sperm cells. Our findings showed that the straight line velocity, the curvilinear velocity, the angular path velocity, and the amplitude of lateral displacement of sperm cells decreased. Moreover, while the level of superoxide dismutase decreased dose-dependently against the toxicity of TRMP-Ps, no significant change was observed in the levels of malondialdehyde and total glutathione. The 4-h median effective concentrations (EC 50 ) of TRMP-Ps based on mobility parameters of sperm ranged from 0.31 mg/L for reduced straight line velocity of sperm cells to 0.51 mg/L for reduced amplitude of lateral displacement of the spermatozoa head. Therefore, we concluded that TRMP-Ps can be a risk for the reproduction cycle of fish in aquatic environments., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

In the context of evaluating the impact of environmentally friendly and sustainably produced alternative protein sources in fish feed, the present study's aim was to examine the overall physiological stress response in one of the main fish species of European freshwater aquaculture, Oncorhynchus mykiss (rainbow trout), following the partial substitution of fish meal (FM) with a Tenebrio molitor (TM) (yellow mealworm) full-fat meal. In total, 222 rainbow trout individuals (115.2 ± 14.2 g) were allocated randomly into six tanks, three per dietary treatment, and were fed a formulated diet containing 60% yellow mealworm (TM60) compared to a control diet without insect meal (TM0). Both diets contained equal amounts of crude protein, dry matter and, lipid content, while the FM in TM60 was 100 g kg -1 corresponding to the one seventh of the TM0. Heat shock response (HSR), MAPK signalling, cell death pathways (apoptosis and autophagy), antioxidant defence mechanisms, and intermediate metabolism were evaluated. In general, HSR and MAPK signalling were activated in response to the inclusion of T. molitor. Moreover, triggering of apoptotic and autophagic processes and the onset of antioxidant defence mechanisms underlined the existence of physiological stress. Despite the apparent dietary-induced stress, rainbow trout in the present study exhibited no mortality and no significant effects regarding growth performance parameters. Specifically, TM60 dietary inclusion resulted in no changes in final body weight, weight gain, and specific growth rate. However, feed intake depicted a statistically significant decrease in TM60 fish compared to TM0 individuals. Nevertheless, nutrient stress should be considered a limiting factor regarding the utilization of T. molitor in O. mykiss diet due to the associated risks for health and welfare., (© 2024 The Author(s). Journal of Animal Physiology and Animal Nutrition published by Wiley‐VCH GmbH.)

Through emission processes, palladium (Pd) particulates from industrial sources are introduced into a range of ecosystems including freshwater environments. Despite this, research on Pd-induced bioaccumulation, uptake, and toxicity is limited for freshwater fishes. Unlike other metals, there are currently no regulations or protective guidelines to limit Pd release into aquatic systems, indicating a global absence of measures addressing its environmental impact. To assess the olfactory toxicity potential of Pd, the present study aimed to explore Pd accumulation in olfactory tissues, olfactory disruption, and oxidative stress in rainbow trout (Oncorhynchus mykiss) following waterborne Pd exposure. Olfactory sensitivity, measured by electro-olfactography, demonstrated that Pd inhibits multiple pathways of the olfactory system following 96 h of Pd exposure. In this study, the concentrations of Pd for inhibition of olfactory function by 20% (2.5 μg/L; IC20) and 50% (19 μg/L; IC50) were established. Rainbow trout were then exposed to IC20 and IC50 Pd concentrations in combination with varying exposure conditions, as changes in water quality alter the toxicity of metals. Independent to Pd, increased water hardness resulted in decreased olfactory perception owing to ion competition at the olfactory epithelium. No other environmental parameter in this study significantly influenced Pd-induced olfactory toxicity. Membrane-associated Pd was measured at the olfactory rosette and gill following exposure; however, this accumulation did not translate to oxidative stress as measured by the production of malondialdehyde. Our data suggest that Pd is toxic to rainbow trout via waterborne contamination near field-measured levels. This study further demonstrated Pd bioavailability and uptake at water-adjacent tissues, adding to our collective understanding of the toxicological profile of Pd. Taken together, our results provide novel insights into the olfactory toxicity in fish following Pd exposure. Integr Environ Assess Manag 2024;20:1407-1419. © 2024 The Authors. Integrated Environmental Assessment and Management published by Wiley Periodicals LLC on behalf of Society of Environmental Toxicology & Chemistry (SETAC)., (© 2024 The Authors. Integrated Environmental Assessment and Management published by Wiley Periodicals LLC on behalf of Society of Environmental Toxicology & Chemistry (SETAC).)

In recent years, nanocopper (Cu NPs) has gained attention due to its antimicrobial properties and potential for industrial, agricultural, and consumer applications. But it also has several effects on the aquatic environment. Widespread use of various nanoproducts has raised concerns about impacts of different nanoparticle size on environment and biological objects. Spermatozoa is a model for studying the ecotoxic effects of pollutants on cells and organisms. This study aimed to investigate the effects of different sizes of copper nanoparticles on rainbow trout spermatozoa motility, and to compare their effects with copper ionic solution. Computer assisted sperm analysis (CASA) was used to detect movement parameters at activation of gametes (direct effect) with milieu containing nanocopper of primary particle size of 40-60, 60-80 and 100 nm. The effect of the elements ions was also tested using copper sulfate solution. All products was prepared in concentration of 0, 1, 5, 50, 125, 250, 350, 500, 750, and 1000 mg Cu L -1 . Six motility parameters were selected for analysis. The harmful effect of Cu NPS nanoparticle was lower than ionic form of copper but the effect depends on the motility parameters. Ionic form caused complete immobilization (MOT = 0 %, IC 100 ) at 350 mg Cu L -1 whilst Cu NPs solution only decreased the percentage of motile sperm (MOT) up to 76.4 % at highest concentration tested of 1000 mg Cu L -1 of 40-60 nm NPs. Cu NPs of smaller particles size had more deleterious effect than the bigger one particularly in percentage of MOT and for curvilinear velocity (VCL). Moreover, nanoparticles decrease motility duration (MD). This may influence fertility because the first two parameters positively correlate with fertilization rate. However, the ionic form of copper has deleterious effect on the percentage of MOT and linearity (LIN), but in some concentrations it slightly increases VCL and MD., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

An in vitro study using rainbow trout spermatozoa was designed to evaluate the toxic effects of different concentrations of captan (CPT), mancozeb (MCZ), and azoxystrobin (AZX) fungicides on motility parameters, lipid peroxidation, SOD activity, total antioxidant capacity (TAC), and DPPH inhibition. Moreover, changes in fatty acids profiles caused by the fungicides were determined for the first time. The results revealed that motility parameters, SOD activities, TAC values, and DPPH inhibitions decreased significantly while lipid peroxidation increased after ≥2 µg/L of CPT, ≥1 µg/L of MCZ, and ≥5 µg/L of AZX incubations for 2 h at 4 °C. Additionally, 10 µg/L CPT, 5 µg/L MCZ, and 200 µg/L AZX reduced motility to the 50 % level. Our results clearly demonstrated significant changes in the fatty acids profiles of spermatozoa exposed to these concentrations of the fungicides. The highest lipid peroxidation and the lowest monounsaturated and polyunsaturated saturated fatty acids (MUFA and PUFA, respectively) were detected in AZX. Even though the susceptibility of spermatozoa to oxidative damage is generally attributed to PUFA contents, the results of this study have represented that MUFA content could play a part in this tendency. Moreover, the lower concentration of MCZ reduced motility to the % 50 level while it deteriorated the fatty acids profile less than did AZX. Overall, the present study demonstrated that the detrimental effects of the fungicides on mitochondrial respiration and related enzymes have more priority than oxidative stress in terms of their toxicities on spermatozoa. It has also been suggested that fish spermatozoa are a good model for determining changes in the fatty acid profiles by fungicides, probably, by other pesticides and environmental contaminants as well., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

Complex hydrodynamics abound in natural streams, yet the selective pressures these impose upon different size classes of fish are not well understood. Attached vortices are produced by relatively large objects that block freestream flow, which fish routinely utilize for flow refuging. To test how flow refuging and the potential harvesting of energy (as seen in Kármán gaiting) vary across size classes in rainbow trout (Oncorhynchus mykiss; fingerling, 8 cm; parr, 14 cm; adult, 22 cm; n=4 per size class), we used a water flume (4100 l; freestream flow at 65 cm s-1) and created vortices using 45 deg wing dams of varying size (small, 15 cm; medium, 31 cm; large, 48 cm). We monitored microhabitat selection and swimming kinematics of individual trout and measured the flow field in the wake of wing dams using time-resolved particle image velocimetry (PIV). Trout of each size class preferentially swam in vortices rather than the freestream, but the capacity to flow refuge varied according to the ratio of vortex width to fish length (WV:LF). Consistent refuging behavior was exhibited when WV:LF≥1.5. All size classes exhibited increased wavelength and Strouhal number and decreased tailbeat frequency within vortices compared with freestream, suggesting that swimming in vortices requires less power output. In 17% of the trials, fish preferentially swam in a manner that suggests energy harvesting from the shear layer. Our results can inform efforts toward riparian restoration and fishway design to improve salmonid conservation., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2024. Published by The Company of Biologists Ltd.)

The contamination of marine and freshwater environments by nanoplastics is considered a global threat for aquatic biota. Taking into account the most recent concentration range estimates reported globally and recognizing a knowledge gap in polystyrene nanoplastics (PS-NPs) ecotoxicology, the present work investigated the harmful effects of 20 nm and 80 nm PS-NPs, at increasing biological complexity, on the rainbow trout Oncorhynchus mykiss RTG-2 and gilthead seabream Sparus aurata SAF-1 cell lines. Twenty nm PS-NPs exerted a greater cytotoxicity than 80 nm ones and SAF-1 were approximately 4-fold more vulnerable to PS-NPs than RTG-2. The engagement of PS-NPs with plasma membranes was accompanied by discernible uptake patterns and morphological alterations along with a nuclear translocation already within a 30-min exposure. Cells were structurally damaged only by the 20 nm PS-NPs in a time-dependent manner as indicated by distinctive features of the execution phase of the apoptotic cell death mechanism such as cell shrinkage, plasma membrane blebbing, translocation of phosphatidylserine to the outer leaflet of the cell membrane and DNA fragmentation. At last, functional analyses unveiled marked transcriptional impairment at both sublethal and lethal doses of 20 nm PS-NPs, with the latter impacting the "Steroid biosynthesis", "TGF-beta signaling pathway", "ECM-receptor interaction", "Focal adhesion", "Regulation of actin cytoskeleton" and "Protein processing in endoplasmic reticulum" pathways. Overall, a distinct ecotoxicological hazard of PS-NPs at environmentally relevant concentrations was thoroughly characterized on two piscine cell lines. The effects were demonstrated to depend on size, exposure time and model, emphasizing the need for a comparative evaluation of endpoints between freshwater and marine ecosystems., Competing Interests: Declaration of competing interest The authors declare no competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Traditional monoculture systems often face challenges related to solid waste production and nutrient use. This study addresses these concerns by investigating the impact of bicultural farming of burbot (Lota lota) and rainbow trout (Oncorhynchus mykiss) in a recirculating aquaculture system (RAS) and the subsequent alteration of production of solid waste, growth and physiological status. The rationale behind incorporating burbot lies in its potential as a supplementary species to improve overall system efficiency and sustainability. The experimental groups in triplicate represented the different ratios of rainbow trout (T) and burbot (B) in the stock: T70/B30, T85/B15 and T100/B0. Burbot, although not assessed in monoculture due to its limited commercial significance, was incorporated into the study as a supplementary species, expected to enhance the nutrient utilisation through its bottom-feeding behaviour and anticipated consumption of solid waste produced by trout. After 77 days of culture, the survival rates of trout remained consistent across experimental groups, averaging over 99%, while burbot exhibited comparable survival rates despite lower cumulative survival, averaging 88%. Feed conversion ratios showed no significant differences between the groups, indicating consistent feed utilisation. A significantly higher specific growth rate (SGR) in trout was observed in group T70/B30. The SGR values of burbot were marginally low and without significant differences between groups. Among biochemical markers of blood plasma, phosphorus concentrations were significantly higher in group T70/B30 for both trout and burbot, suggesting better access to the feed for both species. Elevated antioxidant activity and evidence of oxidative stress were found for both species in intestinal tissue. The presence of burbot in stock significantly affected the production of suspended solids per gram of applied feed. Therefore, group T100/B0 demonstrated the highest solid production at multiple time points, suggesting a relationship between burbot presence and the production of suspended solids. Starting at the 9-week, the trout monoculture group exhibited significantly higher phosphorus levels in sediments compared to group T70/B30, emphasising the role of burbot in shaping sediment nutrient dynamics in RAS, such as supplementary cleaning fish. After 11 weeks, group T100/B0 displayed significantly higher values of produced suspended solids and their composition: DM, organic DM, and phosphorus content per gram of feed applied. These results provide evidence of the influence of burbot on suspended solids production and characteristics. In conclusion, this study indicates the positive effects of burbot presence on solid production and sediment nutrient composition., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

PFAAs (Perfluoroalkyl acids) are a class of bioaccumulative, persistent and ubiquitous environmental contaminants which primarily occupy the hydrosphere and its sediments. Currently, a paucity of toxicological information exists for short chain PFAAs and complex mixtures. In order to address these knowledge gaps, we performed a 3-week, aqueous exposure of rainbow trout to 3 different concentrations of a PFAA mixture (50, 100 and 500 ng/L) modeled after the composition determined in Lake Ontario. We conducted an additional set of exposures to individual PFAAs (25 nM each of PFOS (12,500 ng/L), PFOA (10,300 ng/L), PFBS (7500 ng/L) or PFBA (5300 ng/L) to evaluate differences in biological response across PFAA congeners. Untargeted proteomics and phosphorylated metabolomics were conducted on the blood plasma and head kidney tissue to evaluate biological response. Plasma proteomic responses to the mixtures revealed several unexpected outcomes including Similar proteomic profiles and biological processes as the PFOS exposure regime while being orders of magnitude lower in concentration and an atypical dose response in terms of the number of significantly altered proteins (FDR < 0.1). Biological pathway analysis revealed the low mixture, medium mixture and PFOS to significantly alter (FDR < 0.05) a number of processes including those involved in lipid metabolism, oxidative stress and the nervous system. We implicate plasma increases in PPARD and PPARG as being directly related to these biological processes as they are known to be important regulators in all 3 processes. In contrast to the blood plasma, the high mixture and PFOA exposure regimes caused the greatest change to the head kidney proteome, altering many proteins being involved in lipid metabolism, oxidative stress and inflammation. Our findings support the pleiotropic effect PFAAs have on aquatic organisms at environmentally relevant doses including those on PPAR signaling, metabolic dysregulation, immunotoxicity and neurotoxicity., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Denina Simmons reports financial support and administrative support were provided by Ontario Tech University. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Crown Copyright © 2024. Published by Elsevier B.V. All rights reserved.)

Cryopreservation of rainbow trout semen under field conditions was analyzed. Straw location over liquid nitrogen level is a crucial variable that affects freezing rate and fertilization yield due to changes in nitrogen vapor external temperature. The objectives were: to analyze cryopreservation protocols by experimentally measuring the cooling rates and fertilization yield of 0.5 ml plastic straws located in nitrogen vapor at different heights corresponding to different external temperatures; to numerically simulate the freezing process, by solving the heat transfer partial differential equations with the corresponding thermo-physical properties of the biological system and the plastic straw; to evaluate and analyze the surface heat transfer coefficient (h) during the freezing process of the straws; to introduce a new variable, the characteristic freezing time (tc), that enables comparison between protocols; this variable was defined as the elapsed period between the initial freezing temperature and a final reference temperature of -40 °C (temperature in which more than 80 % of the water is in a frozen state). The mathematical model predicted the temperature distribution inside the straw, showing a low effect of straw plastic materials (polyethylene-terephthalate glycol, polyvinyl-chloride, and polypropylene) on freezing rates. The average h value obtained from numerical simulations was 25.5 W/m 2 K, close to that obtained from the analytical Nusselt correlation for natural convection. An improvement on fertilization trials was observed when the average external nitrogen temperature was -129.6 °C (temperature range: -94 to -171 °C) with an average tc of 56.8 s (ranging between 47 and 72 s). These results corresponded to a height above the level of liquid nitrogen of 2 cm. Comparison with literature reported data showed satisfactory results. Applying mathematical models in the cryobiology field achieved results that are relevant for cryopreservation activities., Competing Interests: Declaration of competing interest Authors declare no conflict of interest in the present study., (Copyright © 2024 Society for Cryobiology. Published by Elsevier Inc. All rights reserved.)

Supplementing a fishmeal-free diet with yeast extract improves rainbow trout (Oncorhynchus mykiss) growth performance and modulates the hepatic and intestinal transcriptomic response. These effects are often observed in the long term but are not well documented after short periods of fasting. Fasting for a few days is a common practice in fish farming, especially before handling the fish, such as for short sorting, tank transfers, and vaccinations. In the present study, rainbow trout were subjected to a 4-day fast and then refed, for 8 days, a conventional diet containing fishmeal (control diet) or alternative diets composed of terrestrial animal by-products supplemented or not with a yeast extract. During the refeeding period alone, most of the parameters considered did not differ significantly in response to the different feeds. Only the expression of claudin-15 was upregulated in fish fed the yeast-supplemented diet compared to the control diet. Conversely, fasting followed by refeeding significantly influenced most of the parameters analyzed. In the proximal intestine, the surface area of villi significantly increased, and the density of goblet cell tended to decrease during refeeding. Although no distinct plasma immune response or major signs of gut inflammation were observed, some genes involved in the structure, complement pathway, antiviral functions, coagulation, and endoplasmic reticulum stress response of the liver and intestine were significantly regulated by refeeding after fasting. These results indicate that short-term fasting, as commonly practiced in fish farming, significantly alters the physiology of the liver and intestine regardless of the composition of the diet., (© 2024. The Author(s).)

More than a decade of study since the personality pace-of-life syndrome (POLS) hypotheses were first proposed, there is little support for it within species. Lack of experimental control, insufficient sampling in the face of highly labile behavioural and metabolic traits, and context dependency of trait correlations are suggested as reasons. Here, I argue that artificial selection and/or use of existing selected lines represents a powerful but under-used approach to furthering our understanding of the POLS. To illustrate this potential, I conducted a focussed review of studies that compared the behaviour, metabolism, growth and survival of an artificially selected fast-growing rainbow trout relative to wild unselected strains, under varying food and risk conditions in the laboratory and field. Resting metabolic rate, food intake, and behaviours that enhance feeding but increase energy expenditure (activity, aggression, boldness), were all higher in the fast strain in paired contrasts, under all food and risk conditions, both in the laboratory and the field. Fast-strain fish grew faster in almost every food and risk situation except where food was highly limited (or absent), had higher survival under low or zero predation risk, but had lower survival under high risk. Several other traits rarely considered in POLS studies were also higher in the fast strain, including maximum swimming speed, and hormones (growth hormone (GH), thyroid hormone (T3) and insulin-like growth factor (IGF-1)). I conclude: (i) assumptions and predictions of the POLS hypothesis are well supported, and (ii) context-dependency was largely absent, but when present revealed trade-offs between food acquisition and predation risk. This focused review highlights the potential of artificial selection in testing POLS ideas, and will hopefully motivate further studies using other animals.

Coldwater species are challenged with increasing water temperatures and fluctuations over their upper thermal limits. This study evaluated the potential of acclimation to higher temperature and dietary antioxidants capacity to mitigate the adverse effects of heat shocks in rainbow trout. To this end, rainbow trout fingerlings were acclimated at optimal (14 °C) and high (20 °C) temperatures and fed on selenium (5 mg/kg) and polyphenol (2 g/kg) supplemented diets for 60 days and then were exposed to heat shocks by increasing water temperature up to 30 °C. Growth performance, survival rate, haemato-immunological parameters, and expression of HSP70α, HSP70β, HSP90β, and IL-1β genes were measured to evaluate the hypothesises. The rainbow trout acclimated to 20 °C and fed on antioxidants supplemented diets showed a significantly higher aftershock survival rate. Moreover, fish acclimated to higher temperature showed higher red blood cell counts as well as serum total protein and albumin during the acclimation trial and heat shocks phase. Acclimation to higher temperature and feeding on antioxidants remarkably enhanced fish immune and antioxidant capacity in comparison to fish adapted to cold water and fed on the basal diet measured by improved respiratory burst and lysozyme activities and upregulation of IL-1β expression during exposure of fish to heat shocks. Furthermore, fish acclimated to higher temperature, especially those fed on antioxidant supplemented diets, showed lower expression levels of HSPs genes during the heat shock phase, indicating that high heat shocks were less stressful for these fish in comparison to cold water acclimated fish. This finding was also supported by lower cortisol levels during heat shocks in fish acclimated to higher temperature. In conclusion, the results of this study indicated that acclimation to higher temperature and/or fed on diets supplemented by selenium and polyphenol, can help to mitigate the adverse effects of the heat shock in rainbow trout., (© 2024. The Author(s).)

Ammonia pollution is an important environmental stress factors in water eutrophication. The intrinsic effects of ammonia stress on liver toxicity and muscle quality of rainbow trout were still unclear. In this study, we focused on investigating difference in muscle metabolism caused by metabolism disorder of rainbow trout liver at exposure times of 0, 3, 6, 9 h at 30 mg/L concentrations. Liver transcriptomic analysis revealed that short-term (3 h) ammonia stress inhibited carbohydrate metabolism and glycerophospholipid production but long-term (9 h) ammonia stress inhibited the biosynthesis and degradation of fatty acids, activated pyrimidine metabolism and mismatch repair, lead to DNA strand breakage and cell death, and ultimately caused liver damage. Metabolomic analysis of muscle revealed that ammonia stress promoted the reaction of glutamic acid and ammonia to synthesize glutamine to alleviate ammonia toxicity, and long-term (9 h) ammonia stress inhibited urea cycle, hindering the alleviation of ammonia toxicity. Moreover, it accelerated the consumption of flavor amino acids such as arginine and aspartic acid, and increased the accumulation of bitter substances (xanthine) and odorous substances (histamine). These findings provide valuable insights into the potential risks and hazards of ammonia in eutrophic water bodies subject to rainbow trout., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Background: In this study, we investigated the effects of swimming activity and feed restriction on digestion and antioxidant enzyme activities in juvenile rainbow trout (average body weight of 26.54 ± 0.36 g)., Methods: The stomach, liver and kidney tissues were obtained from four distinct groups: the static water group (fish were kept in static water and fed to satiation), the feeding restricted group (fish were kept in static water with a 25% feed restriction), the swimming exercised group (fish were forced to swimming at a flow rate of 1 Body Length per second (BL/s)) and the swimming exercised-feed restricted group (subjected to swimming exercise at a 1 BL/s flow rate along with a 25% feed restriction). We determined the levels of glutathione, lipid peroxidation and the activities of catalase, superoxide dismutase, glutathione peroxidase, glutathione reductase, glucose-6-phosphate dehydrogenase and lactate dehydrogenase, as well as the presence of reactive oxygen species in the tissues obtained from the fish. Additionally, the activities of pepsin, protease, lipase and arginase in these tissues were measured., Results: Swimming activity and feed restriction showed different effects on the enzyme activities of the fish in the experimental groups., Conclusion: It can be concluded that proper nutrition and exercise positively influence the antioxidant system and enzyme activities in fish, reducing the formation of free radicals. This situation is likely to contribute to the fish's development., (© 2024 The Authors. Veterinary Medicine and Science published by John Wiley & Sons Ltd.)

In this study, different amounts of glutamine were added to the diet of rainbow trout, and they were then fed for a period of 90 days. The current research investigated the effects of glutamine on various aspects of rainbow trout, including growth performance, condition factor, viscerosomatic index, hepatosomatic index, carcass composition, fatty acid profile, hematological parameters, and biochemical parameters. The study's findings revealed that adding glutamine to the diet of rainbow trout had a beneficial impact on their growth features. The rainbow trout group that was fed a 2% concentration of glutamine experienced the most notable increase in growth rate. A statistically significant difference in growth was observed among all groups (p < 0.05). Adding glutamine to the diet increased the amount of protein and decreased the fat content in the flesh of the fish. Glutamine exerted an influence on the blood and biochemistry parameters of fish, as well as their fatty acid composition. In conclusion, the inclusion of glutamine in the diet of fish could potentially enhance their immune system, improve the quality of their muscles, and enhance their growth performance., (© 2024 The Authors. Journal of Fish Biology published by John Wiley & Sons Ltd on behalf of Fisheries Society of the British Isles.)

This study investigated the effects of occupational enrichment, specifically underwater currents, on the stress status of rainbow trout (Oncorhynchus mykiss). A total of 540 fish were divided into three groups: control tanks without artificial currents (CO), tanks with randomly fired underwater currents (RFC), and tanks with continuous current throughout the day (CT). After 30 days, half of the fish in each group underwent a 5-day pre-slaughter fasting (5D), while the others were fed until the day before slaughter (0D). Fish in the RFC group exhibited lower levels of plasma cortisol and acetylcholinesterase enzyme activity in hypothalamus and optic tract than other groups, suggesting an improved stress status. RFC group also showed higher levels of non-esterified fatty acids (NEFA) in 5D fish and higher liver glycogen stores, suggesting improved energy reserves. In comparison, the CT group had higher LDH levels, possibly due to their increased swimming activity. The CO group had significantly lower NEFA levels at 5D compared to the RFC group, suggesting lower energy reserves. The RFC fish had darker and yellow-reddish skin and liver color, suggesting an improved stress status and lower lipid reserves, respectively. Overall, although a significant stress response was not observed in fasted individuals, possibly due to the relatively short fasting period, the study suggests that providing occupational enrichment using randomly fired underwater currents for 1 month helped to improve stress status in rainbow trout, indicating that occupational enrichment during the grow-out phase can positively impact the welfare of rainbow trout during routine handling procedures., (© 2023. The Author(s), under exclusive licence to Springer Nature B.V.)

N-(1,3-dimethylbutyl)-N'-phenyl-p-phenylenediamine quinone (6PPDQ) has attracted significant attention due to its highly acute lethality to sensitive salmonids. However, studies investigating the mechanisms underlying its acute toxicity have been lacking. In this work, we demonstrated the sensitivity of rainbow trout to 6PPDQ-induced mortality. Moribund trout exhibited significantly higher brain concentrations of 6PPDQ compared to surviving trout. In an in vitro model using human brain microvascular endothelial cells, 6PPDQ can penetrate the blood-brain barrier and enhance blood-brain barrier permeability without compromising cell viability. The time spent in the top of the tank increased with rising 6PPDQ concentrations, as indicated by locomotion behavior tests. Furthermore, 6PPDQ influenced neurotransmitter levels and mRNA expression of neurotransmission-related genes in the brain and exhibited strong binding affinity to target neurotransmission-related proteins using computational simulations. The integrated biomarker response value associated with neurotoxicity showed a positive linear correlation with trout mortality. These findings significantly contribute to filling the knowledge gap between neurological impairments and apical outcomes, including behavioral effects and mortality, induced by 6PPDQ., Competing Interests: Conflict of interest The authors declare that they have no conflict of interest., (Copyright © 2023 Science China Press. Published by Elsevier B.V. All rights reserved.)

Cognitive enrichment is a promising but understudied type of environmental enrichment that aims to stimulate the cognitive abilities of animals by providing them with more opportunities to interact with (namely, to predict events than can occur) and to control their environment. In a previous study, we highlighted that farmed rainbow trout can predict daily feedings after two weeks of conditioning, the highest conditioned response being elicited by the combination of both temporal and signalled predictability. In the present study, we tested the feeding predictability that elicited the highest conditioned response in rainbow trout (both temporal and signalled by bubbles, BUBBLE + TIME treatment) as a cognitive enrichment strategy to improve their welfare. We thus analysed the long-term effects of this feeding predictability condition as compared with an unpredictable feeding condition (RANDOM treatment) on the welfare of rainbow trout, including the markers in the modulation of brain function, through a multidisciplinary approach. To reveal the brain regulatory pathways and networks involved in the long-term effects of feeding predictability, we measured gene markers of cerebral activity and plasticity, neurotransmitter pathways and physiological status of fish (oxidative stress, inflammatory status, cell type and stress status). After almost three months under these predictability conditions of feeding, we found clear evidence of improved welfare in fish from BUBBLE + TIME treatment. Feeding predictability allowed for a food anticipatory activity and resulted in fewer aggressive behaviours, burst of accelerations, and jumps before mealtime. BUBBLE + TIME fish were also less active between meals, which is in line with the observed decreased expression of transcripts related to the dopaminergic system. BUBBLE + TIME fish tented to present fewer eroded dorsal fin and infections to the pathogen Flavobacterium psychrophilum. Decreased expression of most of the studied mRNA involved in oxidative stress and immune responses confirm these tendencies else suggesting a strong role of feeding predictability on fish health status and that RANDOM fish may have undergone chronic stress. Fish emotional reactivity while isolated in a novel-tank as measured by fear behaviour and plasma cortisol levels were similar between the two treatments, as well as fish weight and size. To conclude, signalled combined with temporal predictability of feeding appears to be a promising approach of cognitive enrichment to protect brain function via the physiological status of farmed rainbow trout in the long term., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Pectoral fins play a crucial role in fish locomotion. Despite fishes living in complex fluid environments that exist in rivers and tidal flows, the role of the pectoral fins in navigating turbulent flows is not well understood. This study investigated the kinematics and muscle activity of pectoral fins in rainbow trout as they held station in the unsteady flows behind a D-section cylinder. We observed two distinct pectoral fin behaviors, one during braking and the other during Kármán gaiting. These behaviors were correlated to whole-body movements in response to the hydrodynamic conditions of specific regions in the cylinder wake. Sustained fin extensions during braking, where the fin was held out to maintain its position away from the body and against the flow, were associated with the cessation of forward body velocity, where the fish avoided the suction region directly downstream of the cylinder. Transient fin extensions and retractions during Kármán gaiting controlled body movements in the cross-stream direction. These two fin behaviors had different patterns of muscle activity. All braking events required recruitment from both the abductor and adductor musculature to actively extend a pectoral fin. In contrast, over 50% of fin extension movements during Kármán gaiting proceed in the absence of muscle activity. We reveal that in unsteady fluid environments, pectoral fin movements are the result of a complex combination of passive and active mechanisms that deviate substantially from canonical labriform locomotion, the implications of which await further work on the integration of sensory and motor systems., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2024. Published by The Company of Biologists Ltd.)

Postprandial kinetics of genes expression of gastric (chitinase, pepsinogen) and intestinal (alkaline phosphatase, maltase) digestive enzymes and nutrient transporters (peptide transporter 1, sodium-glucose transporter 1), Brush Border Membrane (BBM) enzymes activity (alkaline phosphatase, leucine aminopeptidase, maltase, saccharase) and blood biochemistry (triglycerides, cholesterol, protein, albumin, glucose, amino acids) through NMR spectroscopy, were investigated in rainbow trout (Oncorhynchus mykiss) fed a commercial aquafeed. For this purpose, fish were starved 72 h and digestive tract and blood were sampled before the meal and at 1.5, 3, 6, 9, 12, and 24 h after feeding (T0, T1.5, T3, T6, T9, T12 and T24). The postprandial kinetic showed that the expression of the genes involved in digestion and nutrient transport, the activity of BBM enzymes, and the presence of metabolites in blood were stimulated in different ways by the presence of feed in the digestive tract. The expression of most genes peaked 3 h after meal except gastric pepsinogen and maltase in distal intestine that peaked at T9 and T12, respectively. The activity of BBM enzymes were stimulated differently based on the intestine tract. The plasma proteins level increased from T1.5 until T9, while the other blood parameters unvariated during the postprandial period. This study supplied useful information about the physiological effects a single meal as a potential tool for planning nutritional studies involving the digestive functions., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Using dominance hierarchies in juvenile rainbow trout (Oncorhynchus mykiss) as a model of chronic social stress in fish, we explored whether epigenetic transcriptional and post-transcriptional mechanisms are involved in the gene expression of corticotropin-releasing factor (crf) and 11β-hydroxysteroid dehydrogenase (11βhsd2), key factors involved in the regulation of the endocrine stress axis response. In juvenile rainbow trout pairs, subordinate individuals display sustained elevation of circulating cortisol concentrations. Cortisol production is controlled by the hypothalamic-pituitary-interrenal (HPI) axis in fish and initiated by CRF release from the preoptic area (POA). Given that crf is modulated during chronic social stress, and that such stress has been implicated in the epigenetic regulation of crf in other taxa, we probed a role for epigenetic regulation of crf transcript abundance in chronically stressed rainbow trout. We also investigated the regulation of the cortisol-metabolising enzyme 11βhsd2 in the POA, which is upregulated in subordinates. The potential involvement of DNA methylation and microRNAs (miRNAs) in the regulation of crf transcript abundance was investigated during social stress in the POA of fish, as was the potential involvement of miRNAs in 11βhsd2 regulation. Although transcript abundances of crf were elevated in subordinate fish after 4 days, DNA methylation profiles within putative promoter sequences upstream of the crf gene were not significantly affected by chronic stress. An inverse relationship between crf and its predicted posttranscriptional regulator miR-103a-3p in the POA suggests that miRNAs may be involved in mediating the effects of chronic social stress on key components of the endocrine stress axis., Competing Interests: Declaration of Competing Interest The authors declare no competing interests., (Copyright © 2023 Elsevier Inc. All rights reserved.)

A new "less invasive" device incorporating an ultrasonic flow probe and a divided chamber, but no stitching of membranes to the fish, was employed to make the first direct measurements of ventilatory flow rate (V̇w) and % O 2 utilization (%U) in juvenile rainbow trout (37 g, 8ºC) after exhaustive exercise (10-min chasing) and voluntary feeding (2.72% body mass ration). Under resting conditions, the allometrically scaled V̇w (300 ml kg -1  min -1 for a 37-g trout = 147 ml kg -1  min -1 for a 236-g trout exhibiting the same mass-specific O 2 consumption rate, ṀO 2 ) and the convection requirement for O 2 (CR = 4.13 L mmol -1 ) were considerably lower, and the %U (67%) was considerably higher than in previous studies using surgically attached masks or the Fick principle. After exhaustive exercise, V̇w and ṀO 2 approximately doubled whereas frequency (fr) and %U barely changed, so increased ventilatory stroke volume (Vsv) was the most important contributor to increased ṀO 2 . CR declined slightly. Values gradually returned to control conditions after 2-3 h. After voluntary feeding, short-term increases in V̇w, Vsv and ṀO 2 were comparable to those after exercise, and fr again did not change. However, %U increased so CR declined even more. The initial peaks in V̇w, Vsv and ṀO 2, similar to those after exercise, were likely influenced by the excitement and exercise component of voluntary feeding. However, in contrast to post-exercise fish, post-prandial fish exhibited second peaks in these same parameters at 1-3 h after feeding, and %U increased further, surpassing 85%, reflecting the true "specific dynamic action" response. We conclude that respiration in trout is much more efficient than previously believed., (© 2023. Crown.)

Described as the 'main ecological factor', temperature, strongly affects the physiological stress responses of fish. In order to evaluate the effects of temperature variations on fish culture and food value chain, the present study was designed as a climate change model. Furthermore, the present study provides a theoretical basis for a better understanding of the mechanisms of the environmentally induced changes. In this direction, we examined the blood physiology and oxidative stress responses induced by temperature variation in the rainbow trout, a temperature-sensitive cold-water fish. The obtained results showed that climate changes promoted the inhibited activities' expressions and the development of potential tissue and hematological defense mechanisms against temperature-induced toxic damage. This study showed that climate change could be a subset of the studies on the stress physiology in aquaculture, which can be developed for new experimental designs and research collaborations. Furthermore, it highlights knowledge gaps to guide future research in this emerging field., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Alak et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

The present study was designed to assess the influence of dietary supplementation with chestnut bee pollen at various levels in rainbow trout, Oncorhynchus mykiss. For two weeks feeding period, a total of 300 fish were allocated into 12 fiberglass tanks and divided into four equal groups, three replicates each, with chestnut bee pollen (BP) dietary inclusion as follows; the fish group was given a basal diet (C); fish group fed a diet supplemented with BP 1% (BP-1); fish group fed a diet supplemented with BP 2% (BP-2); and fish group fed a diet supplemented with BP 4% (BP-3). At the end of the experiment, growth, haematological values, immune status, antioxidant status, and survival rate against Aeromonas salmonicida subsp. achromogenes were evaluated. Dietary supplementation with chestnut bee pollen significantly improves growth performance. Fish fed the diets containing chestnut bee pollen had higher the haematological values than those fed the control diet. The results showed that all the immunological parameters in the groups fed with chestnut bee pollen were significantly higher when compared to the control group. Moreover, dietary chestnut bee pollen increased disease resistance against Aeromonas salmonicida subsp. achromogenes compared to the control group. The tissue SOD, CAT and GSH-Px activities of groups fed with chestnut bee pollen significantly enhanced when compared with the control groups. In contrast, the tissue MDA levels in all groups fed with chestnut bee pollen were significantly decreased. The best values for the antioxidant parameters were determined in the groups fed with 2 and 4% of chestnut bee pollen. Overall, these findings suggest that dietary chestnut bee pollen enhances the growth, the haematological values, the immune and antioxidant response and increases disease resistance against rainbow trout.

Muscle shortening underpins most skeletal motion and ultimately animal performance. Most animal muscle generates its greatest mechanical output over a small, homogeneous range of shortening magnitudes and speeds. However, homogeneous muscle shortening is difficult to achieve for swimming fish because the whole body deforms like a bending beam: as the vertebral column flexes laterally, longitudinal muscle strain increases along a medio-lateral gradient. Similar dorsoventral strain gradients have been identified as the vertebral column flexes dorsally during feeding in at least one body location in one fish. If fish bodies also deform like beams during dorsoventral feeding motions, this would suggest the dorsal body (epaxial) muscles must homogenize both dorsoventral and mediolateral strain gradients. We tested this hypothesis by measuring curvature of the anterior vertebral column with XROMM and muscle shortening in 14 epaxial subregions with fluoromicrometry during feeding in rainbow trout (Oncorhynchus mykiss). We compared measured strain with the predicted strain based on beam theory's curvature-strain relationship. Trout flexed the vertebrae dorsally and laterally during feeding strikes, yet when flexion in both planes was included, the strain predicted by beam theory was strongly and significantly correlated with measured strain (P<0.01, R2=0.60). Beam theory accurately predicted strain (slope=1.15, compared with ideal slope=1) across most muscle subregions, confirming that epaxial muscles experience dorsoventral and mediolateral gradients in longitudinal strain. Establishing this deformation-curvature relationship is a crucial step to understanding how these muscles overcome orthogonal strain gradients to produce powerful feeding and swimming behaviours., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2023. Published by The Company of Biologists Ltd.)

Natural substances has been identified to maintain health and improve growth performance in the aquaculture. The effect of Origanum onites on growth and immune response of rainbow trout was investigated. Experimental groups (A and B) of 70 fish were separated into 10 different treatments. A groups were fed with dietary administration of O. onites essential oil (0.5 mL kg-1 and 3.0 mL kg-1) and crude powder (1.0 g kg-1 and 10.0 g kg-1) for a period of 8 weeks. Other groups (B) were vaccinated against Yersinia ruckeri at the beginning of experiment and then fed the same diets described above. Results showed that feed conversion ratio in fish fed a combination of O. onites and vaccine was statistically better than the control. NBT-positive cells, phagocytic activity, serum lysozyme activity and immunoglobulin M level were stimulated in both non vaccinated and vaccinated fish (p<0.05). Cumulative mortality in fish fed O. onites was lower than controls following challenge with Y. ruckeri. No mortality was observed in vaccinated fish fed with 0.5 mL kg-1 of O. onites. These results indicated that dietary administration of O. onites could act as an enhanced non specific immune response, growth performance and resistance to Y. ruckeri.

The Rainbow trout gill cell-line (RTgill-W1) has been accepted by the Organisation for Economic Co-operation and Development (OECD TG249) as a replacement for fish in acute toxicity tests. In these tests cells are exposed under static conditions. In contrast, in vivo, water moves over fish gills generating fluid shear stress (FSS) that alters cell physiology and response to toxicants. The current study uses a specialised 3D printed chamber designed to house inserts and allows for the flow (0.2 dynes cm 2 ) of water over the cells. This system was used to assess RTgill-W1 cell responses to FSS in the absence and presence of copper (Cu) over 24 h. FSS caused increased gene expression of mechanosensitive channel peizo1 and the Cu-transporter atp7a, elevated reactive oxygen species generation and increased expression of superoxidase dismutase. Cell metabolism was unaffected by Cu (0.163 μM to 2.6 μM Cu) under static conditions but significantly reduced by FSS + Cu above 1.3 μM. Differential expression of metallothionein (mt) a and b was observed with increased expression of mta under static conditions and mtb under FSS on exposure to Cu. These findings highlight toxicologically relevant mechanosensory responses by RTgill-W1 to FSS that may influence toxicological responses., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

The accumulation and tissue distribution of toxicants in aquatic biota can be determinative of their toxic impact to both exposed organisms and their potential human consumers. In the present study, accumulation of the trace metal thallium (Tl) in gill, muscle, plasma, and otoliths of rainbow trout (Oncorhynchus mykiss) following acute (96-h) and subchronic (28-day) waterborne exposures was investigated. Owing to known interactions between Tl and potassium ions (K + ), plasma and muscle K + concentrations were also determined. Branchial Tl accumulated in a dose-dependent manner in both acute and subchronic exposures, while plasma Tl was rapidly mobilized to tissues and accumulated only at exposure concentrations of 141 µg L -1 or higher. For muscle tissue, Tl concentrations at 28 days were markedly lower than those at 96 h at comparable exposure concentrations (0.9 µg L -1 ), indicating the presence of mechanisms that act to reduce Tl accumulation over time. However, after acute exposure, muscle Tl reached concentrations that, if consumed, would exceed acceptable daily intake values for this element, indicating some risk to human health from the consumption of fish from waters heavily contaminated with Tl. Otoliths showed Tl concentrations that reflected exposure concentration and length, confirming their capacity to provide insight into fish exposure history. No changes in tissue K + concentrations were observed, suggesting that accumulation of Tl in rainbow trout plasma and muscle does not occur at the expense of K + homeostasis. In addition to highlighting the capacity of rainbow trout to accumulate Tl to levels that exceed recommended dietary doses to human consumers, the present study provides the first data of tissue-specific Tl accumulation in an important regulatory species. Environ Toxicol Chem 2023;42:1553-1563. © 2023 SETAC., (© 2023 SETAC.)

Rainbow trout (Oncorhynchus mykiss) is traditionally considered as a poor user of digestible carbohydrates harbouring persistent postprandial hyperglycaemia and decreased growth performances when fed a diet containing more than 20% of digestible carbohydrates. While this glucose-intolerant phenotype is well-described in juveniles, evidence points to a particular regulation of glucose metabolism in rainbow trout broodstrocks. By detecting changes in glucose levels and triggering a specific metabolic response, the hypothalamus plays a key role in the regulation of peripheral glucose metabolism. Therefore, our objective was to assess, for the first time in fish, the short-term consequences in hypothalamus, the glucose sensing and feed intake regulation of feeding mature female and male, and neomale rainbow trout with a diet containing either no or a 33% carbohydrate. The hypothalamic glucosensing capacity was assessed through mRNA levels of glucosensing related-genes and feed intake regulation through appetite-regulating peptides. Our data indicate that a brief period of carbohydrate intake (5 meals at 8 °C) did not induce specific changes in glucosensing capacity and appetite-regulating peptides in the hypothalamus of rainbow trout broodstock. Our results did however demonstrate, for the first time in fish, the existence of sex dimorphism of glucosensing-related genes and appetite-regulating peptides., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Cortisol is a key stress-related hormone involved in the physiological adjustments of fish. In gills, cortisol contributes to acclimatization to changes in environmental salinity, promoting both ion uptake or salt excretion. Cortisol exerts its biological effects through its interaction with specific intracellular glucocorticoid (GR) and mineralocorticoid (MR) receptors. Additionally, the further identification of GR and MR on the surface of different tissues, together with the existence of cortisol-mediated effects observed using membrane-impermeable analogs (e.g., cortisol-BSA), supports the existence of membrane-initiated cortisol actions in fish. Nevertheless, the impact of this alternative cortisol mechanism in relevant tissues for fish salinity acclimation, such as gill, is unknown. In this work, we sought to explore the contribution of rapid membrane-initiated cortisol on GR and MR regulation in rainbow trout (Oncorhynchus mykiss) gills using in vivo and in vitro approaches. Juvenile rainbow trout intraperitoneally injected with cortisol or cortisol-BSA showed increased gr2 but no gr1 or mr mRNA levels in gills after one hour of treatment. This result was further confirmed using RT-gills-W1 cell lines stimulated with both versions of cortisol. Interestingly, after three and six hours of cortisol or cortisol-BSA treatment, there were no changes in the mRNA levels of any corticosteroid receptor in RT-gills-W1 cells. Finally, using immunofluorescence analysis, we identified GR and MR in rainbow trout gill cells localized on the cell surface. Considering the in vivo and in vitro results of this work, we suggest that membrane-initiated cortisol action contributes to the early expression of gr2 in rainbow trout gills during salinity acclimation., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Inc. All rights reserved.)

The present study aimed to determine the effects of dietary probiotic supplementation on feed efficiency, physiological parameters, and semen quality of male rainbow trout (Oncorhynchus mykiss) broodstock. For this purpose, a total of 48 breeders with an average initial weight of 1366.1 ± 33.8 g were divided into 4 groups and 3 replicates. Fish were fed with diets containing 0 (control), 1 × 10 9 (P1), 2 × 10 9 (P2), and 4 × 10 9 (P3) CFU multi-strain probiotic kg -1 diet for 8 weeks. According to the results, P2 treatment significantly enhanced body weight increase, specific growth rate, and protein efficiency ratio and decreased feed conversion ratio. Moreover, the highest values of red blood cells count, hemoglobin, and hematocrit values were observed in P2 treatment (P < 0.05). The lowest levels of glucose, cholesterol, and triglyceride were found in P1, P2, and P3 treatments, respectively. Also, the highest levels of total protein and albumin were obtained in P2 and P1 treatments (P < 0.05). Based on the results, plasma enzymes contents were significantly decreased in P2 and P3 treatments. In terms of immune parameters, the complement component 3, complement component 4, and immunoglobulin M levels were increased in all probiotic-fed treatments (P < 0.05). For spermatological features, the highest spermatocrit value, sperm concentration, and motility time were observed in the P2 treatment (P < 0.05). Consequently, we conclude that multi-strain probiotics can be used as functional feed additives in male rainbow trout broodstock to enhance semen quality, improve physiological responses, and better feed efficiency., (© 2023. The Author(s), under exclusive licence to Springer Nature B.V.)

In this study, the effects of temperature on hatching, yolk-sac absorption, larval metamorphosis, post-metamorphic growth, developmental morphology, and muscle cellularity were assessed in rainbow trout, during its early development (until 52 days post-hatching, dph). From the eyed-ova stage, embryos were exposed to either low (8 ± 1 °C, LT-8) or high (16 ± 1 °C, HT-16) temperatures until hatching. Following hatching, half of the sac-fry from LT-8 group were shifted to higher temperature (16 ± 1 °C, LHT-16), and half from HT-16 group were shifted to medium temperature (13 ± 1 °C, HMT-13), for larval rearing. Incubating the eyed-ova at 16 °C preceded the hatching by 6 days, synchronized hatching duration, and minimized hatchlings' size-variation. However, it yielded smaller and morphologically less developed individuals compared to those incubated continuously at 8 ± 1 °C. Post-hatch shifting of sac-fry to high and medium temperatures, respectively, from the initial low and high regimes differentially affected the length and weight of fish. The effect on length was immediate and temporary, but on weight, it appeared to be permanent. Red muscle hypertrophy was observed to be high in HT-16 and HMT-13 individuals (high-temperature incubated groups). White muscle hypertrophy was high in HT-16 and LHT-16 individuals (high post-hatch rearing temperature groups). The effect of early-life temperature regimes on developmental morphology was found to be strong at 22 dph (82.5%) and comparatively weak at 52 dph (65%). The post-hatch rearing temperature caused an immediate but temporary effect on fin development, mainly pectoral, caudal, and anal fin (seen only at 22 dph, not at 52 dph). Contrarily, incubation temperature affected fin position, in a delayed but persistent manner (subtle at 22 dph, but stronger at 52 dph). Overall, this study provides new insights on temperature-dependent changes in developmental morphology, muscle cellularity, and larval growth in rainbow trout and shows that incubation temperature affects ontogeny profoundly than post-hatch thermal regimes., (© 2023. The Author(s), under exclusive licence to Springer Nature B.V.)

The gastrointestinal tract (GIT) lumen of teleosts harbors extreme conditions, especially after feeding: high PCO 2 (20-115 Torr), total ammonia (415-3710 μM), PNH 3 (79-1760 μTorr in the intestine), and virtual anoxia (PO 2  < 1 Torr). These levels could be dangerous if they were to equilibrate with the bloodstream. Thus, we investigated the potential equilibration of O 2 , CO 2 , and ammonia across the GIT epithelia in freshwater rainbow trout by monitoring postprandial arterial and venous blood gases in vivo and in situ. In vivo blood was sampled from the indwelling catheters in the dorsal aorta (DA) and subintestinal vein (SIV) draining the posterior intestine in the fasting state and at 4 to 48 h following catheter-feeding. To investigate possible ammonia absorption in the anterior part of the GIT, blood was sampled from the DA, SIV and hepatic portal vein (HPV) from anaesthetized fish in situ following voluntary feeding. We found minimal equilibration of all three gases between the GIT lumen and the SIV blood, with the latter maintaining pre-feeding levels (PO 2  = 25-49 Torr, PCO 2  = 6-8 Torr, and total ammonia = 117-134 μM and PNH 3  = 13-30 μTorr at 48 h post-feeding). In contrast to the SIV, we found that the HPV total ammonia more than doubled 24 h after feeding (128 to 297 μM), indicative of absorption in the anterior GIT. Overall, the GIT epithelia of trout, although specialized for absorption, prevent dangerous levels of PO 2 , PCO 2 and ammonia from equilibrating with the blood circulation., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Although the gastrointestinal tract (GIT) is an important site for nitrogen metabolism in teleosts, the mechanisms of ammonia absorption and transport remain to be elucidated. Both protein catabolism in the lumen and the metabolism of the GIT tissues produce ammonia which, in part, enters the portal blood through the anterior region of the GIT. The present study examined the possible roles of different GIT sections of rainbow trout (Oncorhynchus mykiss) in transporting ammonia in its unionized gas form-NH 3 -by changing the PNH 3 gradient across GIT epithelia using in vitro gut sac preparations. We also surveyed messenger RNA expression patterns of three of the identified Rh proteins (Rhbg, Rhcg1, and  Rhcg2) as potential NH 3 transporters and NKCC as a potential ammonium ion (NH 4 + ) transporter along the GIT of rainbow trout. We found that ammonia absorption is not dependent on the PNH 3 gradient despite expression of Rhbg and Rhcg2 in the intestinal tissues, and Rhcg2 in the stomach. We detected no expression of Rhbg in the stomach and no expression of Rhcg1 in any GIT tissues. There was also a lack of correlation between ammonia transport and [NH 4 + ] gradient despite NKCC expression in all GIT tissues. Regardless of PNH 3 gradients, the stomach showed the greatest absorption and net tissue consumption of ammonia. Overall, our findings suggest nitrogen metabolism zonation of GIT, with stomach serving as an important site for the absorption, handling and transport of ammonia that is independent of the PNH 3 gradient., (© 2022 Wiley Periodicals LLC.)

Fish scales have been reported to incorporate cortisol over long periods of time and thus provide a promising means of assessing long-term stress in many species of teleost fish. However, the quantification of other stress related hormones has only been accomplished in our previous study conducted in goldfish (Carassius auratus). DHEA is a precursory androgen with anti-stress effects used alongside cortisol to diagnose chronic stress via the cortisol:DHEA ratio in mammals. Included in DHEA's anti-stress mechanisms are changes in the metabolism of cortisol to its inactive metabolite cortisone suggesting the relationships between cortisol, DHEA and cortisone may be additionally informative in the assessment of long-term stress. Therefore, to further explore these concepts in a native fish species and generate more comprehensive comparisons between scale and serum hormone concentrations than was possible in our previous study we implemented a 14-day stress protocol in adult rainbow trout (Oncorhynchus mykiss) and quantified resulting scale and serum cortisol, cortisone and DHEA concentrations. As predicted, elevations in scale concentrations of all hormones were observed in stressed trout compared to controls but were not reflected in serum samples. Significant differences in the cortisol:DHEA and cortisone:cortisol ratios were also found between control and stressed group scales but not serum. These results suggest not only that scales provide a superior medium for the assessment of long-term stress but also that the addition of scale cortisone and DHEA may provide additional relevant information for such assessments., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Fish mortality is associated with harmful algal blooms, although whether toxicity is related directly to the presence of cyanotoxins or the prevailing water chemistry remains unclear. Similarly, while planktivorous fish may be exposed to toxin through the diet, the hazard posed by waterborne extracellular toxin to carnivorous fish is less well understood. In this study rainbow trout (Oncorhynchus mykiss) were exposed for up to 28 d to waterborne microcystin-LR at nominal concentrations of 1.5 and 50 µg L -1 (measured values 2 and 49 µg L -1 , respectively). The former represents the Canadian drinking water guideline, and the latter an elevated environmental level. This study hypothesised that waterborne toxin exposure would specifically impact gill function, and given the importance of this tissue in freshwater fish ion regulation, effects on plasma ions and branchial ion transporter activity would be observed. Microcystin-LR exposure resulted in a significant and persistent hypocalcaemia at the higher exposure concentration, but plasma sodium and branchial activities of the sodium/potassium ATPase, proton ATPase and calcium ATPase enzymes remained unaffected. An in vitro assessment failed to show any effect of microcystin-LR on branchial calcium ATPase activity even at exposure concentrations as high as 1000 µg L -1 . A transient increase in hepatic alkaline phosphatase activity was also observed at 49 µg L -1 , but there were no effects of toxin exposure on branchial or hepatic lactate dehydrogenase activity. These results suggest that microcystin-LR exposure does not have a general effect on ion regulation, but instead produces a novel and specific impact on calcium metabolism in rainbow trout, although the mechanism underlying this effect remains unknown., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Heat stress-induced intestinal damage is a key event in fish pathology. Nano-selenium (nano-Se) shows remarkably high biological activity and low toxicity, making it an ideal and ecological Se formulation; however, to date, the protective effects of nano-Se against heat stress-induced intestinal injury and pertinent molecular mechanisms remain unknown. Herein, rainbow trout (Oncorhynchus mykiss) were fed either a basal diet or basal diet + 5 mg/kg nano-Se. Samples were collected before (18 °C for 9 days; CG18 and Se18 groups) and after (24 °C for 8 h; CG24 and Se24 groups) heat stress treatment. On heat stress exposure, intestinal villus height, muscularis thickness, and goblet cell number decreased, and expression of tight junction proteins (ZO-1, occludin, and claudin-8d) was downregulated; dietary supplementation with nano-Se alleviated these effects. Furthermore, in the presence of nano-Se, catalase activity was elevated, and expression of diverse heat shock proteins (Hsp70b, Hsp90α, and Hsp30), selenoproteins (Gpx1a, Gpx1b1, and Trx), and anti-inflammatory cytokine (TGF-β) was upregulated. In contrast, nano-Se supplementation significantly alleviated the increase of the expression of pro-inflammatory cytokines (IL-1β and TNF-α) and the malondialdehyde content. We also observed that heat stress markedly increased the relative abundance of Actinobacteria, Firmicutes, Methylobacterium, Akkermansia, and Deinococcus and decreased that of Proteobacteria; nano-Se supplementation restored these changes, making their distribution similar to that in the control group. Overall, our findings suggest that nano-Se plays a protective role against heat stress-induced intestinal damage in rainbow trout by promoting the recovery of antioxidant enzyme activity, enhancing protein repair, alleviating inflammatory responses, and restoring intestinal microbiota composition., Competing Interests: Declaration of competing interest The authors declare that they have no conflict of interest., (Copyright © 2023. Published by Elsevier Ltd.)

The gastrointestinal system of fish reacts rapidly to food deprivation. The relative masses of digestive organs and activities of digestive enzymes decrease within days of fasting. This is believed to be an energy-conserving strategy as the metabolic cost of maintaining digestive capacity is high. Cortisol is known for its role in energy mobilization following stress exposure, and prolonged elevated cortisol levels have been shown to reduce growth rates in fish. Fish experiencing chronic cortisol elevations show structural changes to their digestive tissues and overall reductions in relative digestive tissue masses. In fish fasting for prolonged periods, circulating cortisol levels have been reported to be downregulated, upregulated, or unchanged compared to feeding fish. This study aimed to investigate if RU486 and spironolactone, antagonists of the glucocorticoid receptor (GR), and mineralocorticoid receptor (MR), respectively, alone or in combination affect circulating cortisol levels during prolonged starvation. In addition, we tested the effects of blocking GR and MR, on the down-regulation of relative digestive tissue mass during starvation, and its effects on weight loss. Three treatment groups of rainbow trout were intraperitoneally implanted with either GR, MR, or GR and MR blockers. A fourth group was implanted with cortisol, while a fifth group served as a control. All treatment groups were sampled over a course of four weeks of food deprivation and compared against each other and fed control fish at day 0 of the trial. Starvation for 2 weeks and longer significantly increased circulating cortisol levels in all groups except for the group implanted with GR and MR antagonists. Loss of body mass occurred most rapidly during the first week of starvation. Spironolactone treatment resulted in significantly reduced loss of mass during the first week, however, over the following weeks, no differences in mass loss were observed in the groups implanted with blockers, while cortisol-treated fish showed the highest decrease in body mass over time. Relative digestive tissue mass decreased in all groups but apparently, the fasting-induced elevation in plasma cortisol levels did not affect the relative weight loss of digestive tissues as no differences were observed between control fish and GR + MR antagonist treated fish. Very high cortisol levels caused by cortisol treatment however caused a faster decrease in the relative mass of some digestive organs, particularly the stomach., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

The mechanisms involved in hedonic regulation of food intake, including endocannabinoid system (ECs) are scarcely known in fish. We recently demonstrate in rainbow trout the presence of a rewarding response mediated by ECs in hypothalamus and telencephalon when fish fed a lipid-enriched diet, and that central administration of main agonists of ECs namely AEA or 2-AG exert a bimodal effect on feed intake in fish with low doses inducing an increase that disappears with the high dose of both endocannabinoids (EC). To assess the precise involvement of the different receptors of the ECs (CNR1, TRPV1, and GPR55) in this response we injected intracerebroventricularly AEA or 2-AG in the absence/presence of specific receptor antagonists (AM251, capsazepine, and ML193; respectively). The presence of antagonists clearly counteracts the effect of EC supporting the specificity of EC action inducing changes not only in ECs but also in GABA and glutamate metabolism ultimately leading to the increase observed in food intake response., Competing Interests: Declaration of competing interest The authors declare no competing or financial interests., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Environmental condition, such as environmental complexity or stocking density, can directly or indirectly influence animal emotion and ultimately, affective state. Affective states of animals can be assessed through judgement bias tests, evaluating responses to ambiguous situations. In this study, we aimed to determine whether environmental complexity and stocking density impacted rainbow trout affective state. Rainbow trout (n = 108) were housed in recirculating aquaculture systems under commercial conditions while trained at tank-level to discriminate between a positively reinforced chamber (feed) in one location and a negative chamber (positive punishment; chase by net for 1 s) in the opposing location. Fish from successful tanks (two out of five tanks) were then housed in treatment tanks of either high- or low- environmental complexity at either high (165 fish/m 3 ) or low (69 fish/m 3 ) stocking density. Trained fish were tested for latencies to approach three intermediate, ambiguous chambers. Fish housed in high-density tanks were faster to enter all chambers than those housed in low-density tanks (8.5 s vs. 15.2 s; P = 0.001), with faster entries into the positive (7.4 s vs. 15.2 s; P = 0.02) and near-negative chambers (10.2 s vs. 17.4 s; P = 0.006), suggesting that these fish were more optimistic to receive a feed reward. Tank complexity did not affect test outcomes. No differences between treatments were observed between body weight, length, and plasma cortisol. Overall, rainbow trout are capable of discriminating between cues during a judgement bias test and fish housed in high-density environments respond more optimistically in ambiguous situations compared to fish in low-density environments., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Acute warming raises the metabolism of fish, which is matched by increased heart rate. However, thermal acclimation may reduce heart rate through combinations of lowered intrinsic pacemaker rate and increased inhibitory vagal tone. This could affect the baroreflex, which regulates arterial blood pressure through heart rate changes via altered vagal tone. Using pharmacological tools, we assessed autonomic tones and baroreflex regulation of heart rate in rainbow trout (Oncorhynchus mykiss) at 11 °C, and after acute (24 h, 17°C acute ) or chronic (>7 weeks, 17°C chronic ) warming to 17 °C. We hypothesised that warm acclimation would manifest as reduced heart rate and elevated vagal tone in 17°C chronic trout relative to 11 °C and 17°C acute trout, which would increase baroreflex gain and the scope for f H increase through vagal release during hypotension. Compared to 11 °C, the 17°C acute group exhibited slightly higher heart rate (Q 10  = 1.5) and a strong trend for elevated vagal tone (54%). Surprisingly, however, routine heart rate was unaltered by warm acclimation (Q 10  = 1.6), while intrinsic heart rate and vagal tone (22%) declined. Consequently, baroreflex sensitivity to reduced blood pressure was elevated in the 17°C acute group but returned towards 11 °C conditions in the 17°C chronic group. Atropine abolished nearly all chronotropic changes. Bradycardic responses to hypertension and cardiac adrenergic tone were unaltered across temperature treatments. The lack of a clear acclimation effect on routine heart rate in the present study is likely explained by a seasonal effect as the experiments were performed in early winter. Nonetheless, we conclude that baroreflex sensitivity in trout is thermally plastic; the heightened baroreflex sensitivity to hypotension following acute warming likely serves to safeguard tissue oxygen delivery as metabolism is elevated, while the reduced baroreflex sensitivity observed with warm acclimation may be linked to a pronounced metabolic down-regulation., Competing Interests: Declaration of competing interest The authors declare no competing interests., (Copyright © 2022 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Rainbow trout (Oncorhynchus mykiss) is a typical cold-water aquaculture fish and a high-end aquatic product. When water temperature exceeds its optimal range of 12-18 °C, the immune system of rainbow trout becomes weakened and unbalanced. High temperature in summer and global warming severely impact rainbow trout industry. The focus of this study was to explore the mechanisms regulating the immune response of rainbow trout under high temperature stress and identify molecular elements that account for resistance to high temperature. In this study, individual fish were screened in a high temperature stress experiment and divided into resistant (R) and sensitive (S) groups. The hepatic transcriptome sequencing and analysis of mRNAs and microRNAs of the R, S, and control groups showed that the number of the differentially expressed genes (DEGs) in the S group (9259) was higher than that in the R group (5313). Furthermore, the 1233 genes differentially expressed between S and R groups were mainly enriched in immune-related pathways, including cytokine-cytokine receptor interaction, TNF signaling and IL-17 signaling. Among these DEGs were miR-301b-5p and its target gene that encodes nuclear factor of activated T cells two interacting protein (nfatc2ip). The dual-luciferase reporter system and immunofluorescence experiments verified the relationship between miR-301b-5p and nfatc2ip. We also showed that expression levels of miR-301b-5p and nfatc2ip significantly negatively correlated in the liver of rainbow trout under high temperature stress. By performing functional experiments, we showed that activation of miR-301b-5p expression or inhibition of nfatc2ip expression stimulated the phosphorylation of p65, p38, and JNK in the classical nuclear factor kappa-B and mitogen-activated protein kinase pathways under high temperature stress. These manipulations initially promoted the secretion of the pro-inflammatory factor IL-1β and then increased the levels of IL-6, IL-12, and TNF-α. In addition, activation of miR-301b-5p expression or inhibition of nfatc2ip expression stimulated the repair of the hepatic ultrastructural damage caused by high temperature stress by activating the inflammatory response in rainbow trout liver., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Occupational enrichment emerges as a promising strategy for improving the welfare of farmed animals. This form of enrichment aims to stimulate cognitive abilities of animals by providing them with more opportunities to interact with and control their environment. Predictability of salient daily events, and in particular predictability of feeding, is currently one of the most studied occupational enrichment strategies and can take several forms. In fish, while temporal predictability of feeding has been widely investigated, signalled predictability (based on a signal, such as light or sound) has received little attention. Depending on the type of predictability used and the ecology of the species, the effects on fish welfare often differ. The present study aimed to determine which feeding predictability would be most appropriate for rainbow trout, the main continental farmed fish in Europe, and what the consequences might be for their welfare. We tested four feeding predictability conditions: temporal (based on time of day), signalled (based on bubble diffusion), temporal + signalled (based on time and bubble diffusion), and unpredictable (random feeding times). Behavioural and zootechnical outcomes recorded were swimming activity, aggressive behaviours, burst of accelerations, and jumps, emotional reactivity, and growth. Our results showed that rainbow trout can predict daily feedings relying on time and/or bubbles as predictors as early as two weeks of conditioning, as evidenced by their increased swimming activity before feeding or during feed omission tests, which allowed to reinforce their conditioned response. Temporal predictability alone resulted in an increase in pre-feeding aggressive behaviours, burst of accelerations, and jumps, suggesting that the use of time as the sole predictor of feedings in husbandry practices may be detrimental to fish welfare. Signalled predictability with bubbles alone resulted in fewer pre-feeding agonistic behaviours, burst of accelerations, and jumps than in the temporal predictability condition. The combination of temporal and signalled predictability elicited the highest conditioned response and the level of pre-feeding aggression behaviours, burst of accelerations and jumps tended to be lower than for temporal predictability alone. Interestingly, fish swimming activity during bubble diffusion also revealed that bubbles were highly attractive regardless of the condition. Rainbow trout growth and emotional reactivity were not affected by the predictability condition. We conclude, therefore, that the use of bubbles as a feeding predictor could represent an interesting approach to improve rainbow trout welfare in farms, by acting as both an occupational and physical enrichment., (© 2022. The Author(s).)

This study was performed to investigate the tissue-specific effects of clothianidin on Oncorhynchus mykiss by evaluating the biochemical and histological alterations following 21 days of treatment to environmentally relevant concentrations of 3, 15, and 30 µg/L. The emerged behavioral changes in feeding and swimming performance were considered as adaptive responses to avoid the chemical. The toxic effect of pesticide on nervous system and osmoregulation was evidenced with the inhibition of AChE and Na + K + -ATPase. The sustained lipid peroxidation, ranging from muscle (196%) > brain (154%) > gill (140%) > kidney (129%), might be suggested as a mechanism mediating the inhibition of membrane-bound enzymes. Histological evaluation showed clothianidin-induced lesions appearing as necrosis, atrophy, and edema in muscle, hyperplasia, and hypertrophy causing shortening and fusion of the secondary lamellae in gill, vacuolization, and hydropic degeneration in brain, degeneration of tubular epithelium, and existence of melanomacrophage centers in kidney. The pronounced degenerative changes observed in gill indicate the vulnerability of tissue possibly due to its role as first contact and entry point for the pesticide. Consequently, clothianidin exerted its toxic effects by altering normal behavior, causing neurotoxicity and disturbing osmoregulation. Moreover, the imposed stress was responded in a tissue-specific manner and histological lesions become more severe with increasing concentration. The findings clearly reveal the potential threat caused by environmentally relevant concentrations of clothianidin to early life stages of fish.