In a marine teleost, the significance of oxygen supply for acute thermal tolerance depends upon the context and the endpoint used.

Eight juvenile European seabass were exposed to two thermal ramping protocols with different levels of aerobic activity and tolerance endpoint: the critical thermal maximum for swimming (CTSmax) while exercising aerobically until fatigue and the critical thermal maximum (CTmax) under static conditions until loss of equilibrium (LOE). In the CTSmax protocol, warming caused a profound increase in the rate of oxygen uptake (MO2), culminating in a gait transition from steady aerobic towards unsteady anaerobic swimming, then fatigue at 30.3±0.4°C (mean±s.e.m.). Gait transition and fatigue presumably indicate an oxygen limitation, which reflects the inability to meet the combined demands of swimming plus warming. The CTmax protocol also elicited an increase in MO2, culminating in LOE at 34.0±0.4°C, which is significantly warmer than fatigue at CTSmax. The maximum MO2 achieved in the CTmax protocol was, however, less than 30% of that achieved in the CTSmax protocol. Therefore, the static CTmax did not exploit full cardiorespiratory capacity for oxygen supply, indicating that LOE was not caused by systemic oxygen limitation. Consequently, systemic oxygen supply can be significant for tolerance of acute warming in seabass but this depends upon the physiological context and the endpoint used.

The ecological relevance of critical thermal maxima methodology for fishes.

Critical thermal maxima methodology (CTM) has been used to infer acute upper thermal tolerance in fishes since the 1950s, yet its ecological relevance remains debated. In this study, the authors synthesize evidence to identify methodological concerns and common misconceptions that have limited the interpretation of critical thermal maximum (CTmax ; value for an individual fish during one trial) in ecological and evolutionary studies of fishes. They identified limitations of, and opportunities for, using CTmax as a metric in experiments, focusing on rates of thermal ramping, acclimation regimes, thermal safety margins, methodological endpoints, links to performance traits and repeatability. Care must be taken when interpreting CTM in ecological contexts, because the protocol was originally designed for ecotoxicological research with standardized methods to facilitate comparisons within study individuals, across species and contexts. CTM can, however, be used in ecological contexts to predict impacts of environmental warming, but only if parameters influencing thermal limits, such as acclimation temperature or rate of thermal ramping, are taken into account. Applications can include mitigating the effects of climate change, informing infrastructure planning or modelling species distribution, adaptation and/or performance in response to climate-related temperature change. The authors' synthesis points to several key directions for future research that will further aid the application and interpretation of CTM data in ecological contexts.

Temperature-dependent metabolic consequences of food deprivation in the European sardine.

Aquatic ecosystems can exhibit seasonal variation in resource availability and animals have evolved to cope with the associated caloric restriction. During winter in the NW Mediterranean Sea, the European sardine Sardina pilchardus naturally experiences caloric restriction owing to a decrease in the diversity and quantity of plankton. However, ongoing global warming has had deleterious effects on plankton communities such that food shortages may occur throughout the year, especially under warm conditions in the summer. We investigated the interactive effects of temperature and food availability on sardine metabolism by continuously monitoring whole-animal respiration of groups of control (fed) and food-deprived sardines over a 60-day experiment in winter (12°C) or summer (20°C) conditions under natural photoperiod. In addition, we measured mitochondrial respiration of red muscle fibres, biometric variables and energy reserves of individuals sampled at 30 and 60 days. This revealed that winter food deprivation elicits energy saving mechanisms at whole animal and cellular levels by maintaining a low metabolism to preserve energy reserves, allowing high levels of survival. By contrast, despite energy saving mechanisms at the mitochondrial level, whole animal metabolic rate was high during food deprivation in summer, causing increased consumption of energy reserves at the muscular level and high mortality after 60 days. Furthermore, a 5-day re-feeding did not improve survival, and mortalities continued, suggesting that long-term food deprivation at high temperatures causes profound stress in sardines that potentially impairs nutrient absorption.

Tolerance of an acute warming challenge declines with body mass in Nile tilapia: evidence of a link to capacity for oxygen uptake.

It has been proposed that larger individuals within fish species may be more sensitive to global warming, as a result of limitations in their capacity to provide oxygen for aerobic metabolic activities. This could affect size distributions of populations in a warmer world but evidence is lacking. In Nile tilapia Oreochromis niloticus (n=18, mass range 21-313 g), capacity to provide oxygen for aerobic activities (aerobic scope) was independent of mass at an acclimation temperature of 26°C. Tolerance of acute warming, however, declined significantly with mass when evaluated as the critical temperature for fatigue from aerobic swimming (CTSmax). The CTSmax protocol challenges a fish to meet the oxygen demands of constant aerobic exercise while their demands for basal metabolism are accelerated by incremental warming, culminating in fatigue. CTSmax elicited pronounced increases in oxygen uptake in the tilapia but the maximum rates achieved prior to fatigue declined very significantly with mass. Mass-related variation in CTSmax and maximum oxygen uptake rates were positively correlated, which may indicate a causal relationship. When fish populations are faced with acute thermal stress, larger individuals may become constrained in their ability to perform aerobic activities at lower temperatures than smaller conspecifics. This could affect survival and fitness of larger fish in a future world with more frequent and extreme heatwaves, with consequences for population productivity.

Body mass and cell size shape the tolerance of fishes to low oxygen in a temperature-dependent manner.

Aerobic metabolism generates 15-20 times more energy (ATP) than anaerobic metabolism, which is crucial in maintaining energy budgets in animals, fueling metabolism, activity, growth and reproduction. For ectothermic water-breathers such as fishes, low dissolved oxygen may limit oxygen uptake and hence aerobic metabolism. Here, we assess, within a phylogenetic context, how abiotic and biotic drivers explain the variation in hypoxia tolerance observed in fishes. To do so, we assembled a database of hypoxia tolerance, measured as critical oxygen tensions (Pcrit ) for 195 fish species. Overall, we found that hypoxia tolerance has a clear phylogenetic signal and is further modulated by temperature, body mass, cell size, salinity and metabolic rate. Marine fishes were more susceptible to hypoxia than freshwater fishes. This pattern is consistent with greater fluctuations in oxygen and temperature in freshwater habitats. Fishes with higher oxygen requirements (e.g. a high metabolic rate relative to body mass) also were more susceptible to hypoxia. We also found evidence that hypoxia and warming can act synergistically, as hypoxia tolerance was generally lower in warmer waters. However, we found significant interactions between temperature and the body and cell size of a fish. Constraints in oxygen uptake related to cellular surface area to volume ratios and effects of viscosity on the thickness of the boundary layers enveloping the gills could explain these thermal dependencies. The lower hypoxia tolerance in warmer waters was particularly pronounced for fishes with larger bodies and larger cell sizes. Previous studies have found a wide diversity in the direction and strength of relationships between Pcrit and body mass. By including interactions with temperature, our study may help resolve these divergent findings, explaining the size dependency of hypoxia tolerance in fish.

Settleable atmospheric particulate matter induces stress and affects the oxygen-carrying capacity and innate immunity in Nile tilapia (Oreochromis niloticus).

Steel industry emissions of atmospheric particulate matter are responsible for air to water cross-contamination, which deposits metal/metalloid contaminants in aquatic ecosystems. This source of contamination has not been considered in most of the environmental monitoring protocols. Settleable atmospheric particulate matter (SePM) collected in an area of steel industry influence was used to analyze the sublethal effects on the hematological and innate immunological variables in Nile tilapia (Oreochromis niloticus) after short-term exposure (96 h). Blood samples were analyzed to evaluate the oxygen-carrying transport capacity, innate immune activity and stress biomarkers after exposure to ecologically relevant concentration of SePM. The exposure reduced blood oxygen-carrying capacity by lessening hematocrit, hemoglobin, erythrocyte, and mean corpuscular hemoglobin concentration. Compensatory increments in mean corpuscular volume and mean corpuscular hemoglobin have also been observed. The contaminant impacted the immune system by reducing the number of leukocytes, thrombocytes, and monocytes, total plasma protein, leukocyte respiratory activity, and by increasing lysozyme concentration. Furthermore, the contaminant caused endocrine stress response, raising plasma cortisol and glucose. Therefore, the alterations caused by SePM threatened the capacity of sustaining aerobic metabolism, impaired the immune system, and changed the energy allocation due to both stress response and immune effect. This may have important implications for the impact of SePM on aquatic ecosystems. Future investigations should assess SePM impact on general physiology and aerobic performance, especially to face common ecological challenges such as hypoxia and sustained swimming. These results point out the need to develop proper protocols to address the air-to-water cross-contamination risks by iron ore processing industries.

Group size, temperature and body size modulate the effects of social hierarchy on basal cortisol levels in fishes.

Social rank in a structured society has been linked to basal levels of glucocorticoids in various species, with dominant individuals generally presenting lower levels than subordinates. The biotic and abiotic factors influencing glucocorticoids levels across social ranks are still, however, unclear in fishes. We investigated the influences of group size, fish size, sex, age, and reproduction type, plus water salinity and temperature, on the basal levels of cortisol, the major stress hormone in fishes. A phylogenetically controlled meta-analysis was performed on data from 72 studies over 22 species of fishes. As expected, dominants generally exhibited lower levels of cortisol than subordinates. More importantly, the strength of the correlation between cortisol and rank was modulated by three main factors, group size, environmental temperature, and fish size. Differences in basal cortisol between dominants and subordinates were significantly greater in small groups (dyadic contexts) when compared to larger groups. Differences between dominants and subordinates were also greater in temperate regions when compared to the tropics, and in species with larger body size. These results provide valuable insights into the links among hierarchy, stress and metabolism in fishes.

Guidelines for reporting methods to estimate metabolic rates by aquatic intermittent-flow respirometry.

Interest in the measurement of metabolic rates is growing rapidly, because of the importance of metabolism in advancing our understanding of organismal physiology, behaviour, evolution and responses to environmental change. The study of metabolism in aquatic animals is undergoing an especially pronounced expansion, with more researchers utilising intermittent-flow respirometry as a research tool than ever before. Aquatic respirometry measures the rate of oxygen uptake as a proxy for metabolic rate, and the intermittent-flow technique has numerous strengths for use with aquatic animals, allowing metabolic rate to be repeatedly estimated on individual animals over several hours or days and during exposure to various conditions or stimuli. There are, however, no published guidelines for the reporting of methodological details when using this method. Here, we provide the first guidelines for reporting intermittent-flow respirometry methods, in the form of a checklist of criteria that we consider to be the minimum required for the interpretation, evaluation and replication of experiments using intermittent-flow respirometry. Furthermore, using a survey of the existing literature, we show that there has been incomplete and inconsistent reporting of methods for intermittent-flow respirometry over the past few decades. Use of the provided checklist of required criteria by researchers when publishing their work should increase consistency of the reporting of methods for studies that use intermittent-flow respirometry. With the steep increase in studies using intermittent-flow respirometry, now is the ideal time to standardise reporting of methods, so that - in the future - data can be properly assessed by other scientists and conservationists.

Use of complex physiological traits as ecotoxicological biomarkers in tropical freshwater fishes.

We review the use of complex physiological traits, of tolerance and performance, as biomarkers of the toxicological effects of contaminants in subtropical and tropical freshwater fishes. Such traits are growing in relevance due to climate change, as exposure to contaminants may influence the capacity of fishes to tolerate and perform in an increasingly stressful environment. We review the evidence that the critical oxygen level, a measure of hypoxia tolerance, provides a valuable biomarker of impacts of diverse classes of contaminants. When coupled with measures of cardiorespiratory variables, it can provide insight into mechanisms of toxicity. The critical thermal maximum, a simple measure of tolerance of acute warming, also provides a valuable biomarker despite a lack of understanding of its mechanistic basis. Its relative ease of application renders it useful in the rapid evaluation of multiple species, and in understanding how the severity of contaminant impacts depends upon prevailing environmental temperature. The critical swimming speed is a measure of exercise performance that is widely used as a biomarker in temperate species but very few studies have been performed on subtropical or tropical fishes. Overall, the review serves to highlight a critical lack of knowledge for subtropical and tropical freshwater fishes. There is a real need to expand the knowledge base and to use physiological biomarkers in support of decision making to manage tropical freshwater fish populations and their habitats, which sustain rich biodiversity but are under relentless anthropogenic pressure.

Is starvation a cause of overmortality of the Mediterranean sardine?

Animal mortality is difficult to observe in marine systems, preventing a mechanistic understanding of major drivers of fish population dynamics. In particular, starvation is known to be a major cause of mortality at larval stages, but adult mortality is often unknown. In this study, we used a laboratory food-deprivation experiment, on wild caught sardine Sardina pilchardus from the Gulf of Lions. This population is interesting because mean individual phenotype shifted around 2008, becoming dominated by small, young individuals in poor body condition, a phenomenon that may result from declines in energy availability. Continuous monitoring of body mass loss and metabolic rate in 78 captive food-deprived individuals revealed that sardines could survive for up to 57 days on body reserves. Sardines submitted to long-term caloric restriction prior to food-deprivation displayed adaptive phenotypic plasticity, reducing metabolic energy expenditure and enduring starvation for longer than sardines that had not been calorie-restricted. Overall, entry into critical fasting phase 3 occurred at a body condition of 0.72. Such a degree of leanness has rarely been observed over 34 years of wild population monitoring. Still, the proportion of sardines below this threshold has doubled since 2008 and is maximal in January and February (the peak of the reproductive season), now reaching almost 10 % of the population at that time. These results indicate that the demographic changes observed in the wild may result in part from starvation-related adult mortality at the end of the winter reproductive period, despite adaptive plastic responses.

Cardiac and behavioural responses to hypoxia and warming in free-swimming gilthead seabream, Sparus aurata.

Gilthead seabream were equipped with intraperitoneal biologging tags to investigate cardiac responses to hypoxia and warming, comparing when fish were either swimming freely in a tank with conspecifics or confined to individual respirometers. After tag implantation under anaesthesia, heart rate (fH) required 60 h to recover to a stable value in a holding tank. Subsequently, when undisturbed under control conditions (normoxia, 21°C), mean fH was always significantly lower in the tank than in the respirometers. In progressive hypoxia (100% to 15% oxygen saturation), mean fH in the tank was significantly lower than in the respirometers at oxygen levels down to 40%, with significant bradycardia in both holding conditions below this level. Simultaneous logging of tri-axial body acceleration revealed that spontaneous activity, inferred as the variance of external acceleration (VARm), was low and invariant in hypoxia. Warming (21 to 31°C) caused progressive tachycardia with no differences in fH between holding conditions. Mean VARm was, however, significantly higher in the tank during warming, with a positive relationship between VARm and fH across all temperatures. Therefore, spontaneous activity contributed to raising fH of fish in the tank during warming. Mean fH in respirometers had a highly significant linear relationship with mean rates of oxygen uptake, considering data from hypoxia and warming together. The high fH of confined seabream indicates that respirometry techniques may bias estimates of metabolic traits in some fishes, and that biologging on free-swimming fish will provide more reliable insight into cardiac and behavioural responses to environmental stressors by fish in their natural environment.

Changes in foraging mode caused by a decline in prey size have major bioenergetic consequences for a small pelagic fish.

Global warming is causing profound modifications of aquatic ecosystems and one major outcome appears to be a decline in adult size of many fish species. Over the last decade, sardine populations in the Gulf of Lions (NW Mediterranean Sea) have shown severe declines in body size and condition as well as disappearance of the oldest individuals, which could not be related to overfishing, predation pressure or epizootic diseases. In this study, we investigated whether this situation reflects a bottom-up phenomenon caused by reduced size and availability of prey that could lead to energetic constraints. We fed captive sardines with food items of two different sizes eliciting a change in feeding mode (filter-feeding on small items and directly capturing larger ones) at two different rations for several months, and then assessed their muscle bioenergetics to test for changes in cellular function. Feeding on smaller items was associated with a decline in body condition, even at high ration, and almost completely inhibited growth by comparison to sardines fed large items at high ration. Sardines fed on small items presented specific mitochondrial adjustments for energy sparing, indicating a major bioenergetic challenge. Moreover, mitochondria from sardines in poor condition had low basal oxidative activity but high efficiency of ATP production. Notably, when body condition was below a threshold value of 1.07, close to the mean observed in the wild, it was directly correlated with basal mitochondrial activity in muscle. The results show a link between whole-animal condition and cellular bioenergetics in the sardine, and reveal physiological consequences of a shift in feeding mode. They demonstrate that filter-feeding on small prey leads to poor growth, even under abundant food and an increase in the efficiency of ATP production. These findings may partially explain the declines in sardine size and condition observed in the wild.

Le changement global entraîne de profondes modifications des écosystèmes aquatiques, l'une des principales étant le déclin de la taille des adultes chez de nombreuses espèces de poissons. Au cours de la dernière décennie, les populations de sardines du Golfe du Lion (Nord-Ouest de la Méditerranée) ont montré une importante diminution de leur taille et de leur condition corporelle ainsi qu'une disparition des individus les plus âgés, qui n'ont pas pu être liées à la surpêche, à la pression de prédation ou aux épizooties. Dans cette étude, nous avons cherché à savoir si cette situation reflète un phénomène ascendant causé par la réduction de la taille et de la disponibilité des proies qui pourrait entraîner des contraintes énergétiques chez la sardine. Nous avons ainsi nourri des sardines captives avec des granulés de deux tailles différentes provoquant un changement de mode d'alimentation (filtration des petits granulés et capture directe des plus gros) et à deux rations différentes pendant plusieurs mois, puis nous avons évalué leur bioénergétique musculaire pour tester les changements au niveau de leur fonction cellulaire. L'alimentation à base de petits granulés a été associée à un déclin de la condition corporelle, même à une ration élevée, et à une croissance quasiment inhibée par rapport aux sardines nourries avec des plus gros granulés à une ration élevée. Les sardines nourries avec des petits granulés ont également présenté des ajustements mitochondriaux spécifiques pour économiser de l'énergie, indiquant un défi bioénergétique majeur. De plus, les mitochondries des sardines en mauvaise condition présentaient une faible activité oxydative basale, mais une efficacité élevée de production d'ATP. Notamment, lorsque la condition corporelle était inférieure à une valeur seuil de 1,07, proche de la moyenne observée dans la nature, elle était directement corrélée à l'activité mitochondriale basale dans le muscle. Ces résultats montrent un lien entre la condition de l'animal entier et la bioénergétique cellulaire chez la sardine, et révèlent les conséquences physiologiques d'un changement de mode d'alimentation. Ils démontrent que le nourrissage via la filtration de petites proies entraîne une faible croissance, même en cas de nourriture abondante, et une augmentation de l'efficacité de la production d'ATP. Ces résultats peuvent expliquer en partie le déclin de la taille et de la condition des sardines observé dans la nature.

An investigation of links between metabolic rate and feed efficiency in European sea bass Dicentrarchus labrax.

Feed efficiency (FE) is the amount of body weight gain for a given feed intake. Improving FE through selective breeding is key for sustainable finfish aquaculture but its evaluation at individual level is technically challenging. We therefore investigated whether individual routine metabolic rate (RMR) was a predictor of individual FE in the European sea bass Dicentrarchus labrax, a major species in European mariculture. The European sea bass has three genetically distinct populations across its geographical range, namely Atlantic (AT), West Mediterranean (WM), and East Mediterranean (EM). We compared FE and RMR of fish from these three populations at 18 or 24 °C. We held 200 fish (62 AT, 66 WM, and 72 EM) in individual aquaria and fed them from ad libitum down to fasting. FI was assessed for an ad libitum feeding rate and for a fixed restricted ration (1% of metabolic body weight·day-1, with metabolic body weight = body weight0.8). After being refed 12 wk in a common tank, individual RMR was measured over 36 h by intermittent flow respirometry. There was a significant effect of temperature whereby fish at 18 °C had greater mean FE (P < 0.05) and lower RMR (P < 0.001). There was also a significant effect of population, where AT fish had lower FE (P < 0.05) and greater RMR (P < 0.001) than WM and EM, at both temperatures. Despite these differences in temperature and population means, individual FE and RMR were not significantly correlated (P > 0.05). Therefore, although the results provide evidence of an association between metabolic rate and FE, RMR was not a predictor of individual FE, for reasons that require further investigation.

The role of mechanistic physiology in investigating impacts of global warming on fishes.

Warming of aquatic environments as a result of climate change is already having measurable impacts on fishes, manifested as changes in phenology, range shifts and reductions in body size. Understanding the physiological mechanisms underlying these seemingly universal patterns is crucial if we are to reliably predict the fate of fish populations with future warming. This includes an understanding of mechanisms for acute thermal tolerance, as extreme heatwaves may be a major driver of observed effects. The hypothesis of gill oxygen limitation (GOL) is claimed to explain asymptotic fish growth, and why some fish species are decreasing in size with warming; but its underlying assumptions conflict with established knowledge and direct mechanistic evidence is lacking. The hypothesis of oxygen- and capacity-limited thermal tolerance (OCLTT) has stimulated a wave of research into the role of oxygen supply capacity and thermal performance curves for aerobic scope, but results vary greatly between species, indicating that it is unlikely to be a universal mechanism. As thermal performance curves remain important for incorporating physiological tolerance into models, we discuss potentially fruitful alternatives to aerobic scope, notably specific dynamic action and growth rate. We consider the limitations of estimating acute thermal tolerance by a single rapid measure whose mechanism of action is not known. We emphasise the continued importance of experimental physiology, particularly in advancing our understanding of underlying mechanisms, but also the challenge of making this knowledge relevant to the more complex reality.

Oxygen uptake, heart rate and activities of locomotor muscles during a critical swimming speed protocol in the gilthead sea bream Sparus aurata.

Oxygen uptake, heart rate and contraction frequencies of slow oxidative (SO) and fast glycolytic (FG) muscle were measured simultaneously in gilthead seabream Sparus aurata submitted to stepwise increases in current speed in a swimming respirometer. Variation in oxygen uptake was closely related to variation in heart rate, over initial steps these rose in concert with an increase in contraction frequency of SO muscle. There was an asymptote in oxygen uptake and heart rate at high speeds that reflected a transition from exclusive use of aerobic SO muscle to a combination of SO and anaerobic FG muscle, and which preceded fatigue.

Intraspecific variation in tolerance of warming in fishes.

Intraspecific variation in key traits such as tolerance of warming can have profound effects on ecological and evolutionary processes, notably responses to climate change. The empirical evidence for three primary elements of intraspecific variation in tolerance of warming in fishes is reviewed. The first is purely mechanistic that tolerance varies across life stages and as fishes become mature. The limited evidence indicates strongly that this is the case, possibly because of universal physiological principles. The second is intraspecific variation that is because of phenotypic plasticity, also a mechanistic phenomenon that buffers individuals' sensitivity to negative impacts of global warming in their lifetime, or to some extent through epigenetic effects over successive generations. Although the evidence for plasticity in tolerance to warming is extensive, more work is required to understand underlying mechanisms and to reveal whether there are general patterns. The third element is intraspecific variation based on heritable genetic differences in tolerance, which underlies local adaptation and may define long-term adaptability of a species in the face of ongoing global change. There is clear evidence of local adaptation and some evidence of heritability of tolerance to warming, but the knowledge base is limited with detailed information for only a few model or emblematic species. There is also strong evidence of structured variation in tolerance of warming within species, which may have ecological and evolutionary significance irrespective of whether it reflects plasticity or adaptation. Although the overwhelming consensus is that having broader intraspecific variation in tolerance should reduce species vulnerability to impacts of global warming, there are no sufficient data on fishes to provide insights into particular mechanisms by which this may occur.

Social dynamics obscure the effect of temperature on air breathing in Corydoras catfish.

In some fishes, the ability to breathe air has evolved to overcome constraints in hypoxic environments but comes at a cost of increased predation. To reduce this risk, some species perform group air breathing. Temperature may also affect the frequency of air breathing in fishes, but this topic has received relatively little research attention. This study examined how acclimation temperature and acute exposure to hypoxia affected the air-breathing behaviour of a social catfish, the bronze corydoras Corydoras aeneus, and aimed to determine whether individual oxygen demand influenced the behaviour of entire groups. Groups of seven fish were observed in an arena to measure air-breathing frequency of individuals and consequent group air-breathing behaviour, under three oxygen concentrations (100%, 60% and 20% air saturation) and two acclimation temperatures (25 and 30°C). Intermittent flow respirometry was used to estimate oxygen demand of individuals. Increasingly severe hypoxia increased air breathing at the individual and group levels. Although there were minimal differences in air-breathing frequency among individuals in response to an increase in temperature, the effect of temperature that did exist manifested as an increase in group air-breathing frequency at 30°C. Groups that were more socially cohesive during routine activity took more breaths but, in most cases, air breathing among individuals was not temporally clustered. There was no association between an individual's oxygen demand and its air-breathing frequency in a group. For C.aeneus, although air-breathing frequency is influenced by hypoxia, behavioural variation among groups could explain the small overall effect of temperature on group air-breathing frequency.

Using aerobic exercise to evaluate sub-lethal tolerance of acute warming in fishes.

We investigated whether fatigue from sustained aerobic swimming provides a sub-lethal endpoint to define tolerance of acute warming in fishes, as an alternative to loss of equilibrium (LOE) during a critical thermal maximum (CTmax) protocol. Two species were studied, Nile tilapia (Oreochromis niloticus) and pacu (Piaractus mesopotamicus). Each fish underwent an incremental swim test to determine gait transition speed (UGT), where it first engaged the unsteady anaerobic swimming mode that preceded fatigue. After suitable recovery, each fish was exercised at 85% of their own UGT and warmed 1°C every 30 min, to identify the temperature at which they fatigued, denoted as CTswim Fish were also submitted to a standard CTmax, warming at the same rate as CTswim, under static conditions until LOE. All individuals fatigued in CTswim, at a mean temperature approximately 2°C lower than their CTmax Therefore, if exposed to acute warming in the wild, the ability to perform aerobic metabolic work would be constrained at temperatures significantly below those that directly threatened survival. The collapse in performance at CTswim was preceded by a gait transition qualitatively indistinguishable from that during the incremental swim test. This suggests that fatigue in CTswim was linked to an inability to meet the tissue oxygen demands of exercise plus warming. This is consistent with the oxygen and capacity limited thermal tolerance (OCLTT) hypothesis, regarding the mechanism underlying tolerance of warming in fishes. Overall, fatigue at CTswim provides an ecologically relevant sub-lethal threshold that is more sensitive to extreme events than LOE at CTmax.

Interactive effects of mercury exposure and hypoxia on ECG patterns in two Neotropical freshwater fish species: Matrinxã, Brycon amazonicus and traíra, Hoplias malabaricus.

Hypoxia and mercury contamination often co-occur in tropical freshwater ecosystems, but the interactive effects of these two stressors on fish populations are poorly known. The effects of mercury (Hg) on recorded changes in the detailed form of the electrocardiogram (ECG) during exposure to progressive hypoxia were investigated in two Neotropical freshwater fish species, matrinxã, Brycon amazonicus and traíra, Hoplias malabaricus. Matrinxã were exposed to a sublethal concentration of 0.1 mg L-1 of HgCl2 in water for 96 h. Traíra were exposed to dietary doses of Hg by being fed over a period of 30 days with juvenile matrinxãs previously exposed to HgCl2, resulting in a dose of 0.45 mg of total Hg per fish, each 96 h. Both species showed a bradycardia in progressive hypoxia. Hg exposure impaired cardiac electrical excitability, leading to first-degree atrioventricular block, plus profound extension of the ventricular action potential (AP) plateau. Moreover, there was the development of cardiac arrhythmias and anomalies such as occasional absence of QRS complexes, extra systoles, negative Q-, R- and S-waves (QRS complex), and T wave inversion, especially in hypoxia below O2 partial pressures (PO2) of 5.3 kPa. Sub-chronic dietary Hg exposure induced intense bradycardia in normoxia in traira, plus lengthening of ventricular AP duration coupled with prolonged QRS intervals. This indicates slower ventricular AP conduction during ventricular depolarization. Overall, the data indicate that both acute waterborne and sub-chronic dietary exposure (trophic level transfer), at sublethal concentrations of mercury, cause damage in electrical stability and rhythm of the heartbeat, leading to myocardial dysfunction, which is further intensified during hypoxia. These changes could lead to impaired cardiac output, with consequences for swimming ability, foraging capacity, and hence growth and/or reproductive performance.