Does heat tolerance actually predict animals' geographic thermal limits?

The "climate extremes hypothesis" is a major assumption of geographic studies of heat tolerance and climatic vulnerability. However, this assumption remains vastly untested across taxa, and multiple factors may contribute to uncoupling heat tolerance estimates and geographic limits. Our dataset includes 1000 entries of heat tolerance data and maximum temperatures for each species' known geographic limits (hereafter, Tmax). We gathered this information across major animal taxa, including marine fish, terrestrial arthropods, amphibians, non-avian reptiles, birds, and mammals. We first tested if heat tolerance constrains the Tmax of sites where species could be observed. Secondly, we tested if the strength of such restrictions depends on how high Tmax is relative to heat tolerance. Thirdly, we correlated the different estimates of Tmax among them and across species. Restrictions are strong for amphibians, arthropods, and birds but often weak or inconsistent for reptiles and mammals. Marine fish describe a non-linear relationship that contrasts with terrestrial groups. Traditional heat tolerance measures in thermal vulnerability studies, like panting temperatures and the upper set point of preferred temperatures, do not predict Tmax or are inversely correlated to it, respectively. Heat tolerance restricts the geographic warm edges more strongly for species that reach sites with higher Tmax for their heat tolerance. These emerging patterns underline the importance of reliable species' heat tolerance indexes to identify their thermal vulnerability at their warm range edges. Besides, the tight correlations of Tmax estimates across on-land microhabitats support a view of multiple types of thermal challenges simultaneously shaping ranges' warm edges for on-land species. The heterogeneous correlation of Tmax estimates in the ocean supports the view that fish thermoregulation is generally limited, too. We propose new hypotheses to understand thermal restrictions on animal distribution.

Thermal plasticity over a marine-estuarine ecocline can buffer a tropical fish from warming.

Intraspecific variation in thermal tolerance can favor species persistence in a warmer ocean, but is often overlooked in fine-scale studies. Nonetheless, local drivers (e.g. salinity) interact with temperature to shape species' thermal response. Here, we acclimated juveniles of Brazilian silversides Atherinella brasiliensis captured at the limits of a marine-estuarine ecocline under reciprocal-cross conditions, to test for phenotypic plasticity in heat tolerance. We also tested whether silversides acclimated to temperatures predicted for 2100 (+3-4.5 °C). Fish in warm-brackish waters showed higher CTMax (Critical Thermal Maximum) than those in cold-marine conditions, regardless of their origin. Silversides' CTMax reached up to 40.6 °C, but it did not increase after exposure to temperatures predicted for 2100. Lack of acclimation response suggests that silversides heat tolerance has reached a "ceiling", despite thermal plasticity. Our findings show that fine-scale environmental heterogeneity can promote phenotypic plasticity for tropical species, reducing the risk of short-term extirpation.

Consistent predator-prey biomass scaling in complex food webs.

The ratio of predator-to-prey biomass is a key element of trophic structure that is typically investigated from a food chain perspective, ignoring channels of energy transfer (e.g. omnivory) that may govern community structure. Here, we address this shortcoming by characterising the biomass structure of 141 freshwater, marine and terrestrial food webs, spanning a broad gradient in community biomass. We test whether sub-linear scaling between predator and prey biomass (a potential signal of density-dependent processes) emerges within ecosystem types and across levels of biological organisation. We find a consistent, sub-linear scaling pattern whereby predator biomass scales with the total biomass of their prey with a near ¾-power exponent within food webs - i.e. more prey biomass supports proportionally less predator biomass. Across food webs, a similar sub-linear scaling pattern emerges between total predator biomass and the combined biomass of all prey within a food web. These general patterns in trophic structure are compatible with a systematic form of density dependence that holds among complex feeding interactions across levels of organization, irrespective of ecosystem type.

Does Predation Exacerbate the Risk of Endosymbiont Loss in Heat Stressed Hermatypic Corals? Molecular Cues Provide Insights Into Species-Specific Health Outcomes in a Multi-Stressor Ocean.

Ocean warming has been a major driver of coral reef bleaching and mass mortality. Coupled to other biotic pressures, corals' ability for acclimatization and adaptation may become compromised. Here, we tested the combined effects of warming scenarios (26, 30, and 32°C) and predation (wound vs. no wound) in coral health condition (paleness, bleaching, and mortality), cellular stress responses (heat shock protein 70 kDa Hsp70, total ubiquitin Ub, and total antioxidant capacity TAC), and physiological state (integrated biomarker response index, IBR) of seven Scleractinian coral species, after being exposed for 60 days. Results show that although temperature was the main factor driving coral health condition, thermotolerant species (Galaxea fascicularis, Psammocora contigua, and Turbinaria reniformis) displayed increased paleness, bleaching, and mortality in predation treatments at high temperature, whereas thermosensitive species (Acropora tenuis, Echinopora lamellosa, and Montipora capricornis brown and green morphotypes) all died at 32°C, regardless of predation condition. At the molecular level, results show that there were significant main and interactive effects of species, temperature, and predation in the biomarkers assessed. Temperature affected Hsp70, Ub, and TAC, evidencing the role of protein folding and turnover, as well as reactive oxygen species scavenging in heat stress management. Predation increased Hsp70 and Ub, suggesting the activation of the pro-phenoloxidase system and cytokine activity, whereas the combination of both stressors mainly affected TAC during moderate stress and Ub under severe stress, suggesting that redox balance and defense of homeostasis are crucial in tissue repair at high temperature. IBR levels showed an increasing trend at 32°C in predated coral fragments (although non-significant). We conclude that coral responses to the combination of high temperature and predation pressure display high inter-species variability, but these stressors may pose a higher risk of endosymbiont loss, depending on species physiology and stress intensity.

Impacts of dissolved Zn and nanoparticle forms in the fatty acid landscape of Mytilus galloprovincialis.

The introduction of waste containing heavy metals into the marine environment has been increasing for the past few decades, yet there are still several pending questions regarding how it impacts aquatic fauna. This study compared the effects of zinc exposure in its ionic- and nanoparticle forms on the mussel Mytilus galloprovincialis and sampled at different time-points. Zinc accumulation was observable after one week. Exposure to 100 µg L-1 of either form for 28 days also resulted in the higher depletion of fatty acids, lipid peroxidation products accumulation, and changes in the fatty acid profiles. This was also observed for lower concentrations, although to a smaller extent. Given the importance of fatty acids in the marine trophic chains, these zinc-induced alterations have significant potential of introducing negative impacts on the ecosystem and ultimately on human nutrition. Finally, we show that fatty acids may be used as efficient biomarkers of zinc-induced stress.

Conserved fatty acid profiles and lipid metabolic pathways in a tropical reef fish exposed to ocean warming - An adaptation mechanism of tolerant species?

Climate warming is causing rapid spatial expansion of ocean warm pools from equatorial latitudes towards the subtropics. Sedentary coral reef inhabitants in affected areas will thus be trapped in high temperature regimes, which may become the "new normal". In this study, we used clownfish Amphiprion ocellaris as model organism to study reef fish mechanisms of thermal adaptation and determine how high temperature affects multiple lipid aspects that influence physiology and thermal tolerance. We exposed juvenile fish to two different experimental conditions, implemented over 28 days: average tropical water temperatures (26 °C, control) or average warm pool temperatures (30 °C). We then performed several analyses on fish muscle and liver tissues: i) total lipid content (%), ii) lipid peroxides, iii) fatty acid profiles, iv) lipid metabolic pathways, and v) weight as body condition metric. Results showed that lipid storage capacity in A. ocellaris was not affected by elevated temperature, even in the presence of lipid peroxides in both tissues assessed. Additionally, fatty acid profiles were unresponsive to elevated temperature, and lipid metabolic networks were consequently well conserved. Consistent with these results, we did not observe changes in fish weight at elevated temperature. There were, however, differences in fatty acid profiles between tissue types and over time. Liver showed enhanced α-linolenic and linoleic acid metabolism, which is an important pathway in stress response signaling and modulation on environmental changes. Temporal oscillations in fatty acid profiles are most likely related to intrinsic factors such as growth, which leads to the mobilization of energetic reserves between different tissues throughout time according to organism needs. Based on these results, we propose that the stability of fatty acid profiles and lipid metabolic pathways may be an important thermal adaptation feature of fish exposed to warming environments.

Different sensitivity to heatwaves across the life cycle of fish reflects phenotypic adaptation to environmental niche.

Predicting responses of marine organisms to global change requires eco-physiological assessments across the complex life cycles of species. Here, we experimentally tested the vulnerability of a demersal temperate fish (Sparus aurata) to long-lasting heatwaves, on larval, juvenile and adult life-stages. Fish were exposed to simulated coastal (18 °C), estuarine (24 °C) summer temperatures, and heatwave conditions (30 °C) and their physiological responses were assessed based on cellular stress response biomarkers (heat shock protein 70 kDa, ubiquitin, antioxidant enzymes, lipid peroxidation) and phenotypic measures (histopathology, condition and mortality). Life-stage vulnerability can be ranked as larvae > adults > juveniles, based on mortality, tissue pathology and the capacity to employ cellular stress responses, reflecting the different environmental niches of each life stage. While larvae lacked acclimation capacity, which resulted in damage to tissues and elevated mortality, juveniles coped well with elevated temperature. The rapid induction of cytoprotective proteins maintained the integrity of vital organs in juveniles, suggesting adaptive phenotypic plasticity in coastal and estuarine waters. Adults displayed lower plasticity to heatwaves as they transition to deeper habitats for maturation, showing tissue damage in brain, liver and muscle. Life cycle closure of sea breams in coastal habitats will therefore be determined by larval and adult stages.

Engineered metal nanoparticles in the marine environment: A review of the effects on marine fauna.

There is an increasing awareness of how damaging pollutants in the marine environment can be, however information on the effects of metal engineered nanoparticles (ENPs) on marine biota is still insufficient, despite an exponential rising in related publications in recent years. In order to provide an integrated insight on the present state of the art on metal ENP-related ecotoxicology studies on marine fauna, this review aimed to: (i) highlight the means of toxicity of metal ENPs in the marine environment, (ii) identify the principal biotic and abiotic factors that may alter metal ENP toxicity, and (iii) analyse and categorize results of these studies, including accumulation, molecular and histological biomarkers, genotoxicity and behavioural changes. Data retrieved from Scopus yielded 134 studies that met pre-established criteria. Most often, the target ENPs were titanium, zinc, copper or silver, and most studies (61.2%) focused on the phylum Mollusca. The degree of toxicity of metal ENPs was often dependent on the concentrations tested, length of exposure and the type of tissue sampled. Effects from simple tissue accumulation to DNA damage or behavioural alterations were identified, even when concentrations below environmentally available levels were used. It is proposed that other phyla besides the traditional Mollusca (and within it Bivalvia) should be used more often in this kind of studies, that exact pathways of toxicity be further explored, and lastly that co-stressors be used in order to best mimic conditions observed in nature. In this review, the current knowledge on engineered metal nanoparticles and their effects on marine fauna was summarized, highlighting present knowledge gaps. Guidelines for future studies focusing on under-developed subjects in ENP toxicology are also briefly provided.

Long-term exposure to increasing temperatures on scleractinian coral fragments reveals oxidative stress.

Global warming is leading to increases in tropical storms' frequency and intensity, allowing fragmentation of reef-forming coral species, but also to coral bleaching and mortality. The first level of organism's response to an environmental perturbation occurs at the cellular level. This study investigated the long-term oxidative stress on fragments of nine Indo-Pacific reef-forming coral species exposed for 60 days to increasing temperatures (30 °C and 32 °C) and compared results with control temperature (26 °C). Coral overall condition (appearance), lipid peroxidation (LPO), catalase activity (CAT), and glutathione S-transferase (GST) were assessed. The species Turbinaria reniformis, Galaxea fascicularis, and Psammocora contigua were the most resistant to heat stress, presenting no oxidative damage at 30 °C. Unlike G. fasciularis, both T. reniformis and P. contigua showed no evidence of oxidative damage at 32 °C. All remaining species' fragments died at 32 °C. Stylophora pistillata and Pocillopora damicornis were the most susceptible species to heat stress, not resisting at 30 °C.

Predator traits determine food-web architecture across ecosystems.

Predator-prey interactions in natural ecosystems generate complex food webs that have a simple universal body-size architecture where predators are systematically larger than their prey. Food-web theory shows that the highest predator-prey body-mass ratios found in natural food webs may be especially important because they create weak interactions with slow dynamics that stabilize communities against perturbations and maintain ecosystem functioning. Identifying these vital interactions in real communities typically requires arduous identification of interactions in complex food webs. Here, we overcome this obstacle by developing predator-trait models to predict average body-mass ratios based on a database comprising 290 food webs from freshwater, marine and terrestrial ecosystems across all continents. We analysed how species traits constrain body-size architecture by changing the slope of the predator-prey body-mass scaling. Across ecosystems, we found high body-mass ratios for predator groups with specific trait combinations including (1) small vertebrates and (2) large swimming or flying predators. Including the metabolic and movement types of predators increased the accuracy of predicting which species are engaged in high body-mass ratio interactions. We demonstrate that species traits explain striking patterns in the body-size architecture of natural food webs that underpin the stability and functioning of ecosystems, paving the way for community-level management of the most complex natural ecosystems.

Molecular assessment of wild populations in the marine realm: Importance of taxonomic, seasonal and habitat patterns in environmental monitoring.

Scientists are currently faced with the challenge of assessing the effects of anthropogenic stressors on aquatic ecosystems. Cellular stress response (CSR) biomarkers are ubiquitous and phylogenetically conserved among metazoans and have been successfully applied in environmental monitoring but they can also vary according to natural biotic and abiotic factors. The reported variability may thus limit the wide application of biomarkers in monitoring, imposing the need to identify variability levels in the field. Our aim was to carry out a comprehensive in situ assessment of the CSR (heat shock protein 70 kDa, ubiquitin, antioxidant enzymes) and oxidative damage (lipid peroxidation) in wild populations across marine taxa by collecting fish, crustaceans, mollusks and cnidarians during two different seasons (spring and summer) and two habitat types (coast and estuary). CSR end-point patterns were different between taxa with mollusks having higher biomarker levels, followed by the cnidarians, while fish and crustaceans showed lower biomarker levels. The PCA showed clear clusters related to mobility/sessile traits with sessile organisms showing greater levels (>2-fold) of CSR proteins and oxidative damage. Mean intraspecific variability in the CSR measured by the coefficient of variation (% CV) (including data from all seasons and sites) was elevated (35-94%). Overall, there was a seasonal differentiation in biomarker patterns across taxonomic groups, especially evident in fish and cnidarians. A differentiation in biomarker patterns between habitat types was also observed and associated with phenotypic plasticity or local adaptation. Overall, specimens collected in the estuary had lower biomarker levels when compared to specimens collected in the coast. This work highlights the importance of assessing baseline biomarker levels across taxa, seasons and habitats prior to applying biomarker analyses in environmental monitoring. Selecting bioindicator species, defining sampling strategies, and identifying confounding factors are crucial preliminary steps that ensure the success of biomarkers as powerful tools in biomonitoring.

Thermal stress and energy metabolism in two circumtropical decapod crustaceans: Responses to acute temperature events.

Extreme events associated with global warming, such as ocean heat waves, can have contrasting fitness consequences for different species, thereby modifying the structure and composition of marine communities. Here, we examined the effects of a laboratory simulated heat wave on the physiology and performance of two Indo-Pacific crustacean species: the shrimp Rhynchocinetes durbanensis and the hermit crab Calcinus laevimanus. We exposed the crustaceans to a control temperature or to a +5 °C temperature (25 °C vs 30 °C) for two consecutive weeks, and weekly analyzed protective proteins, antioxidant activity, and lipid peroxides in muscle and visceral mass. Fulton's K, total protein, %C, and C:N molar ratio of muscle tissue were also analyzed at the end of the experiment. Results showed that 1) the most responsive tissues were the muscle in the shrimp species and the visceral mass in the hermit crab species; 2) biomarker responses in both species occurred mostly after 7 days of exposure; 3) temperature stress led to an increase in biomarker levels; 4) highest biomarker fold-changes were detected in protective chaperones and antioxidants superoxide dismutase and glutathione-S-transferase; 4) integrated biomarker indices suggested poorer health status in individuals subjected to the heat wave; 5) performance changes at the organism level were only detected in R. durbanensis; and 6) mortality rates of both species remained unchanged with the heat wave. Finally, we concluded that these species are capable of physiological adjustments in response to rapid environmental changes, which ultimately confers them with enough thermal tolerance to withstand this simulated heat wave without major consequences for fitness.

What's in a tide pool? Just as much food web network complexity as in large open ecosystems.

Understanding the fundamental laws that govern complex food web networks over large ecosystems presents high costs and oftentimes unsurmountable logistical challenges. This way, it is crucial to find smaller systems that can be used as proxy food webs. Intertidal rock pool environments harbour particularly high biodiversity over small areas. This study aimed to analyse their food web networks to investigate their potential as proxies of larger ecosystems for food web networks research. Highly resolved food webs were compiled for 116 intertidal rock pools from cold, temperate, subtropical and tropical regions, to ensure a wide representation of environmental variability. The network properties of these food webs were compared to that of estuaries, lakes and rivers, as well as marine and terrestrial ecosystems (46 previously published complex food webs). The intertidal rock pool food webs analysed presented properties that were in the same range as the previously published food webs. The niche model predictive success was remarkably high (73-88%) and similar to that previously found for much larger marine and terrestrial food webs. By using a large-scale sampling effort covering 116 intertidal rock pools in several biogeographic regions, this study showed, for the first time, that intertidal rock pools encompass food webs that share fundamental organizational characteristics with food webs from markedly different, larger, open and abiotically stable ecosystems. As small, self-contained habitats, intertidal rock pools are particularly tractable systems and therefore a large number of food webs can be examined with relatively low sampling effort. This study shows, for the first time that they can be useful models for the understanding of universal processes that regulate the complex network organization of food webs, which are harder or impossible to investigate in larger, open ecosystems, due to high costs and logistical difficulties.

Environmental health assessment of warming coastal ecosystems in the tropics - Application of integrative physiological indices.

According to climate science, ocean warming is one of the current and future greatest threats to coastal ecosystems. Projection scenarios for the end of this century show that tropical intertidal ecosystems are particularly at risk. In this study we optimized and tested a holistic method for bio-monitoring present and projected thermal pressure in such ecosystems, in order to assess organism vulnerability to ocean warming. Several species representative of different animal groups (fish, crustaceans and gastropods) were collected from the field and subjected to an experimental trial for 28 days, testing two temperatures: control (present seawater summer temperature) and elevated temperature (+3 °C, projected seawater temperature anomaly for 2100). Muscle samples were collected weekly to quantify several biomarkers of: i) macromolecular damage (protein unfolding and denaturation, and lipid peroxidation), ii) reactive oxygen species (ROS) scavengers (antioxidant enzymes), and iii) body condition (energy reserves and body mass). These biomarkers were combined in integrated biomarker response (IBR) indices, either in three separate stress response categories (as previously defined) or in a unique combined analysis of overall physiological performance. Both approaches suggest that temperature affected IBRs, with increasing temperatures significantly impairing the overall health of individuals. Biomarkers of lower levels of biological organization indicated deleterious effects of temperature, whereas biomarkers of higher levels suggested maintenance of performance after chronic exposure. Overall indices combining the estimates of biomarkers across levels of biological organization are essential to predict the vulnerability of species, or populations, to climate warming. Such indices may assist managers and stakeholders in the establishment of monitoring programs and environmental policies toward the conservation of fragile coastal systems.

Ecological traps in shallow coastal waters-Potential effect of heat-waves in tropical and temperate organisms.

Mortality of fish has been reported in tide pools during warm days. That means that tide pools are potential ecological traps for coastal organisms, which happen when environmental changes cause maladaptive habitat selection. Heat-waves are predicted to increase in intensity, duration and frequency, making it relevant to investigate the role of tide pools as traps for coastal organisms. However, heat waves can also lead to acclimatization. If organisms undergo acclimatization prior to being trapped in tide pools, their survival chances may increase. Common tide pool species (46 species in total) were collected at a tropical and a temperate area and their upper thermal limits estimated. They were maintained for 10 days at their mean summer sea surface temperature +3°C, mimicking a heat-wave. Their upper thermal limits were estimated again, after this acclimation period, to calculate each species' acclimation response. The upper thermal limits of the organisms were compared to the temperatures attained by tide pool waters to investigate if 1) tide pools could be considered ecological traps and 2) if the increase in upper thermal limits elicited by the acclimation period could make the organisms less vulnerable to this threat. Tropical tide pools were found to be ecological traps for an important number of common coastal species, given that they can attain temperatures higher than the upper thermal limits of most of those species. Tide pools are not ecological traps in temperate zones. Tropical species have higher thermal limits than temperate species, but lower acclimation response, that does not allow them to survive the maximum habitat temperature of tropical tide pools. This way, tropical coastal organisms seem to be, not only more vulnerable to climate warming per se, but also to an increase in the ecological trap effect of tide pools.

Molecular Plasticity under Ocean Warming: Proteomics and Fitness Data Provides Clues for a Better Understanding of the Thermal Tolerance in Fish.

Ocean warming is known to alter the performance and fitness of marine organisms albeit the proteome underpinnings of species thermal tolerance are still largely unknown. In this 1-month experiment we assessed the vulnerability of the gilt-head sea bream Sparus aurata, taken here as a biological model for some key fisheries species, to ocean warming (control 18°C, nursery ground temperature 24°C and heat wave 30°C). Survival was impaired after 28 days, mainly at 30°C although fishes' condition was unaltered. Muscle proteome modulation was assessed at 14 and 21 days, showing that protein expression profiles were similar between fish exposed to 18 and 24°C, differing from fish exposed to 30°C. Fish subjected to 24°C showed an enhanced glycolytic potential and decreased glycogenolysis mainly at 14 days of exposure. Fish subjected to 30°C also showed enhanced glycolytic potential and up-regulated proteins related to gene expression, cellular stress response (CSR), and homeostasis (mostly cytoskeletal dynamics, acid-base balance, chaperoning). However, inflammatory processes were elicited at 21 days along with a down-regulation of the tricarboxylic acid cycle. Thus, juvenile fish seem able to acclimate to 24°C but possibly not to 30°C, which is the predicted temperature for estuaries during heat waves by the year 2100. This may be related with increasing constraints on organism physiology associated with metabolic scope available for performance and fitness at higher temperatures. Consequently, recruitment of commercial sea breams may be in jeopardy, highlighting the need for improved management plans for fish stocks.

Seasonal changes in stress biomarkers of an exotic coastal species - Chaetopleura angulata (Polyplacophora) - Implications for biomonitoring.

Knowledge on baseline values of stress biomarkers in natural conditions is urgent due to the need of reference values for monitoring purposes. Here we assessed the cellular stress response of the chiton Chaetopleura angulata in situ. Biomarkers commonly used in environmental monitoring (heat shock protein 70kDa, total ubiquitin, catalase, glutathione-S-transferase, superoxide-dismutase, lipid peroxidation) were analyzed in the digestive system, gills and muscle of C. angulata, under spring and summer conditions in order to assess seasonal tissue-specific responses. Season had an effect on all targeted organs, especially affecting the digestive system which displayed clear seasonal clusters. The respective Integrated Biomarker Response (IBR) showed a 7.2-fold seasonal difference. Muscle and gills showed similar IBRs between seasons making them appropriate organs to monitor chemical pollution as they were less responsive to seasonal variation. The most stable biomarkers in these organs were ubiquitin and superoxide-dismutase thus being reliable for monitoring purposes.

Comparing biomarker responses during thermal acclimation: A lethal vs non-lethal approach in a tropical reef clownfish.

Knowledge of thermal stress biology for most tropical fish species in reef ecosystems under climate change is still quite limited. Thus, the objective of this study was to measure the time-course changes of thermal stress biomarkers in the commercially exploited coral reef fish Amphiprion ocellaris, during a laboratory simulated event of increased temperature. Heat shock protein 70kDa (Hsp70) and total ubiquitin (Ub) were determined in the muscle (lethal method) and in the fin (non-lethal alternative method) under two temperature treatments (control - 26°C and elevated temperature - 30°C) throughout one month with weekly samplings. Results suggest that biomarker basal levels are tissue-specific and influence the degree of response under temperature exposure. Responses were highly inducible in the muscle but not in fin tissue, indicating that the latter is not reliable for monitoring purposes. Thermal stress was observed in the muscle after one week of exposure (both biomarkers increased significantly) and Ub levels then decreased, suggesting the animals were able to acclimate by maintaining high levels of Hsp70 and through an effective protein turnover. In addition, the results show that mortality rates did not differ between treatments. This indicates that A. ocellaris is capable of displaying a plastic response to elevated temperature by adjusting the protein quality control system to protect cell functions, without decreasing survival. Thus, this coral reef fish species presents a significant acclimation potential under ocean warming scenarios of +4°C. Monitoring of thermal stress through a non-lethal method, fin-clipping, although desirable proved to be inadequate for this species.

Food web of the intertidal rocky shore of the west Portuguese coast - Determined by stable isotope analysis.

The characterization of food web structure, energy pathways and trophic linkages is essential for the understanding of ecosystem functioning. Isotopic analysis was performed on food web components of the rocky intertidal ecosystem in four sites along the Portuguese west coast. The aim was to 1) determine the general food web structure, 2) estimate the trophic level of the dominant organisms and 3) track the incorporation of organic carbon of different origins in the diet of the top consumers. In this food web, fish are top consumers, followed by shrimp. Anemones and gastropods are intermediate consumers, while bivalves and zooplankton are primary consumers. Macroalgae Bifurcaria bifurcata, Ulva lactuca, Fucus vesiculosus, Codium sp. and phytoplankton are the dominant producers. Two energy pathways were identified, pelagic and benthic. Reliance on the benthic energy pathway was high for many of the consumers but not as high as previously observed in subtidal coastal food webs. The maximum TL was 3.3, which is indicative of a relatively short food web. It is argued that the diet of top consumers relies directly on low levels of the food web to a considerable extent, instead of on intermediate levels, which shortens the trophic length of the food web.

Physiological, cellular and biochemical thermal stress response of intertidal shrimps with different vertical distributions: Palaemon elegans and Palaemon serratus.

The ability to cope with high temperature variations is a critical factor in intertidal communities. Two species of intertidal rocky shore shrimps (Palaemon sp.) with different vertical distributions were collected from the Portuguese coast in order to test if they were differentially sensitive to thermal stress. Three distinct levels of biological organization (organismal, biochemical, and cellular) were surveyed. The shrimp were exposed to a constant rate of temperature increase of 1°C x h(-1), starting at 20°C until reaching the CTMax (critical thermal maximum). During heat stress, two biomarkers of protein damage were quantified in the muscle via enzyme-linked immunosorbent assays: heat shock proteins HSP70 (hsp70/hsc70) and total ubiquitin. Muscle histopathological alterations caused by temperature were also evaluated. CTMax values were not significantly different between the congeners (P. elegans 33.4 ± 0.5 °C; P. serratus 33.0 ± 0.5 °C). Biomarker levels did not increase along the temperature trial, but P. elegans (higher intertidal) showed higher amounts of HSP70 and total ubiquitin than P. serratus (lower intertidal). HSP70 and total ubiquitin levels showed a positive significant correlation in both species, suggesting that their association is important in thermal tolerance. Histopathological observations of muscle tissue in P. serratus showed no gross alterations due to temperature but did show localized atrophy of muscle fibers at CTMax. In P. elegans, alterations occurred at a larger scale, showing multiple foci of atrophic muscular fascicles caused by necrotic or autolytic processes. In conclusion, Palaemon congeners displayed different responses to stress at a cellular level, with P. elegans having greater biomarker levels and histopathological alterations.