Groundwater systems of the Yucatan Peninsula form one of the world's most intricate aquifer systems, supporting a unique and ecologically essential subterranean fauna. The physiological capacities of resident species, and their tolerance or ability to cope with changing environmental challenges is unknown for most species. Considering the vertical stratification of anchialine ecosystems, accelerated climate change and anthropogenic pressures, we sought to investigate the physiological characteristics of a key groundwater-restricted species (stygobionts) that is broadly distributed in the Yucatan Peninsula and has been observed in fresh- and marine groundwater. Thus, we (i) characterized the respiratory metabolism, osmoregulatory capacity and thermal tolerance of the cave isopod Creaseriella anops and (ii) evaluated how variations in salinity and oxygen concentration impact its physiological performance and antioxidant defence system. Our findings indicate that C. anops is isosmotic at 17.7‰ (580.8 mOsm/kg) and possesses a strong osmoregulatory capacity. When acclimated to freshwater (0‰) at 26 ± 1°C, C. anops demonstrated a maximum critical temperature of 33.6 ± 1.3°C and a minimum critical temperature of 19.0 ± 2.0°C, with an aerobic scope of 0.053 mg O₂/g/h. Dissolved oxygen levels (15 to 100% saturation) did not influence routine oxygen consumption rates. Acute shifts in salinity (from 0‰ to 8, 14 or 35‰) initially elevated oxygen consumption rates, which returned to routine levels within three hours across all salinity conditions. This metabolic response was associated with a slight activity increase in antioxidant enzymes and elevated protein carbonylation and lipid peroxidation. In summary, C. anops showed outstanding osmoregulatory, metabolic and antioxidant capacities that likely contribute to its wide distribution within the YP's anchialine ecosystems, providing insights into how this species may respond to future environmental shifts.

Antarctic and sub-Antarctic fishes of the genus Harpagifer inhabit extreme environments and face emerging anthropogenic stressors, including plastic pollution and warming. While each factor is known to affect fish physiology, their acute combined impacts on molecular responses and host-microbe interactions remain poorly understood. In this study, we investigated the immediate (24 h) transcriptional response of stress-related genes heat shock protein 70 (hsp70), S100 calcium binding protein (s100), High Mobility Group 1 box (hmg1b), E3 Ubiquitin Ligases (E3), and BCL2 Associated X (Bax) and gut microbiome diversity in H. antarcticus (King George Island, Antarctica) and H. bispinis (Punta Arenas, Chile) exposed to elevated temperature (TI), PVC microplastics (MP), or their combination (TI + MP). Both stressors altered gene transcription in a tissue- and species-specific manner, suggesting a synergistic stress response under combined treatment. Temperature rise consistently modulated stress markers, while acute PVC exposure intensified apoptotic signaling. Notably, species-specific patterns emerged: H. bispinis showed a more pronounced induction of pro-apoptotic pathways, whereas H. antarcticus maintained a higher induction of protective chaperones within this short temporal window. At the microbial level, while community-wide diversity metrics remained statistically stable, an exploratory analysis revealed increased inter-individual variability and the enrichment of specific stress-tolerant bacterial taxa. These results reveal that acute warming and microplastic exposure interactively disturb host, cellular homeostasis, with distinct transcriptional plasticity across closely related species. Our study provides mechanistic insights into the early physiological challenges faced by Notothenioids, highlighting the importance of coupling molecular biomarkers with microbiome analyses to assess the initial stages of response in cold-adapted fishes to environmental change.

Human activities are impacting even the most remote regions of our planet, including Antarctica. The Fildes Bay coastal area (King George Island) is among those considered more at risk to anthropogenic stressors, with increasing concern on consequences to marine life. Here we investigated the potential hazard posed by contaminants of emerging concern (CECs) as nanoplastics and metal-oxide nanomaterials either as single or combined exposure in the Antarctic clam Laternula elliptica. The digestive gland was used to assess the transcriptomic response and oxygen consumption upon in vivo exposure to polystyrene (PS-COOH-NP) and titanium-dioxide nanoparticles (TiO2-NP) at environmental realistic concentrations (5–50 µg · l-1). A total of 2852 differentially expressed genes were found in clams exposed to either single contaminant or in combination versus the control group. The transcript functions affected included autophagy, cell cycle, proteases co-chaperones and their transcription factors, antioxidants, membrane fatty acids, cell pH balance enzymes, mitochondria stress and ATP production in PS-COOH-NP (5 µg · l-1) and TiO2-NP (5 and 50 µg · l-1) groups as well as in co-exposure at 50 µg · l-1. Exposure to TiO2-NP (5 and 50 µg · l-1) and co-exposure at 50 µg · l-1 affected transmembrane transport enzymes and histones. Co-exposure at 5 µg · l-1 shows ribosomal proteins, protease and apoptosis transcripts affected and PS-COOH-NP (at 50 µg · l-1 only) protease as transcripts with known functions. Oxygen consumption decreased significantly upon combined exposure to nano-pollutants at the two concentrations, suggesting a synergistic effect impairing clam metabolism. Our results highlighted several molecular pathways affected by treatments and metabolism decreased in co-exposures, demonstrating the hazardous behavior and toxicity of these nanoparticles.

Barnacle aquaculture is a developing area of production that is currently limited by the reliance on capturing naturally produced larvae. The solution to this limitation would be the development of hatcheries for culturing barnacle larvae and juveniles (Juvs). The giant barnacle Austromegabalanus psittacus (Molina 1788) is an edible species and, thus, of economic importance in Chile. Extensive cultures and hatcheries on an experimental scale have been developed for this species. In this study, a dynamic model, based on data from previously published studies, was developed to evaluate the larval development of A. psittacus over a range of temperatures and salinities in order to identify the optimal culture conditions for obtaining Juvs. The larval development time, survival, physiological, and biochemical responses of A. psittacus larvae were evaluated. The conceptual model representing the dynamic hypothesis, and the simulations, were carried out using Vensim PLE and Stella Architect, respectively. The best production performance under hatchery conditions was obtained at 18 degrees C/32 PSU. Under these conditions, the survival during the development of the nauplius (Nau) stages was 90%, and the development time was 7.5 days. The recently molted cyprids (Cyps) had an accumulated energy of 24.42 J/mg dry weight (dw). At the Cyp stage, the development time was 6 days and the survival was 54%, while the energy of the Juv was 12.02 J/mg dw. Increasing the initial energy of Nau I (NI) resulted in higher survival and energy in the Cyp and Juv stages. These simulations demonstrate the usefulness of these dynamic models in designing and evaluating hatchery protocols that optimize the performance of A. psittacus larvae and may allow scaling to mass cultivation.

Barnacle production is a new aquaculture activity, that has developed to a semi-industrial level with extensive cultures in floating systems. However, the mass culture of larvae and juveniles under controlled hatchery conditions have not yet been developed. In this study, the larval development of the giant barnacle Austromegabalanus psittacus (Molina, 1788) was experimentally examined at different combinations of temperature (9, 12, 15, 18, and 21 degrees C) and salinity (20, 24, 28, and 32 PSU). The development time, mortality and metabolism of the larval stages and their influence on the energy of cyprids and recently metamorphosed juveniles were evaluated. Larval development was completed at 9 and 21 degrees C combined with salinities of 28 and 32 PSU. There was 100 % larval mortality at salinity ranges from 20 to 24 PSU. Larval development time was shorter at 21 degrees C /24 PSU (8.50 f 0.70 days) and longer at 9 degrees C /32 PSU (33.57 f 2.93 days). Oxygen consumption and ammonium excretion increased with the increase in temperature and reduction in salinity. The lowest accumulated mortalities were observed at 18 degrees C with salinities of 28 and 32 PSU, as were the highest content of lipids and proteins, production levels, and net growth efficiency (K2). The results indicate that the combination of temperature and salinity has a synergetic effect on larval development time, survival, and energy accumulation on A. psittacus larvae. These results establish the basis for the design of protocols for cultivating larvae in hatcheries increasing the predictability of juvenile supplies for industrial-scale production of this species.

Invasions by non-indigenous species (NIS) are among the major problems that coastal ecosystems are facing globally and are driven by anthropogenic activities, such as international vessel traffic and aquaculture. Bryozoans are typical constituents of vessel hull-fouling communities, and the phylum contains several notorious globally invasive species. Few NIS, including bryozoan species, are reported along the Chilean Southeast Pacific coast, which has traditionally been attributed to the local oceanographic and ecologic conditions. However, the low numbers may also be the consequence of insufficient monitoring, as well as the small size and morphologic similarity of many species, which hinders their detection. Here, we deployed artificial floating settlement plates in strategic locations (ports/marinas or aquaculture sites) along the Chilean coast (from 29°S to 41°S), to detect non-indigenous bryozoans. With this method, we found a total of 14 species, including five NIS and four cryptogenic species. Additionally, we report two NIS from opportunistic findings in the rocky intertidal zone and from a piece of plastic litter. Three species represent new records from Chile: Bugulina cf. fulva, Bugulina stolonifera, and Watersipora arcuata. These are described in detail in the present paper. The new records importantly enhance the number of known (bryozoan) invasions along the Chilean coast and corroborate the need for regular monitoring. The installation of artificial settlement plates at high-risk sites (especially ports and marinas) has proven to be a successful and efficient low-cost approach, which can help to detect NIS introductions in their early stages, before spreading to natural environments. Regular monitoring of these sites, as well as compulsory legislation for hull cleaning, both for long-distance (nationally and internationally) travelling and local vessels, could greatly reduce the problem.

We examined how a species inhabiting a latitudinal gradient, from warm oxygenated surface waters to cold oxygen-limited subsurface waters along the Eastern South Pacific (ESP) shelf, responds to latitudinal temperature shifts at low-oxygen isopleths. We combined temperature-oxygen sections from the World Ocean Database, historical records of pelagic/benthic Grimothea monodon occurrence across latitude, models with these data, and laboratory experiments assessing juveniles' routine and postprandial metabolism under realistic temperature-oxygen conditions. The life habits (pelagic or benthic) of squat lobsters were related to temperature at the 2 mL O2 L-1 oxygen isopleth. At temperatures > 15 °C near the upper oxygen minimum zone isopleth, mostly pelagic individuals were observed, suggesting restricted vertical migration. The physiological performance of juveniles (main migratory stage) was negatively affected by high temperature-hypoxia interaction. Routine metabolic rates decreased by 60% under hypoxia at 21 °C, and postprandial metabolism (as Specific Dynamic Action) was also strongly reduced under those conditions. Grimothea monodon can shift between pelagic and benthic habitats across a range of ESP conditions, maintaining the intergenerational ability to alternate habitats. This plasticity, expressed as vertical expansion or restriction, may help maintain or expand its latitudinal ranges, with natural food webs and fisheries adjusting to its availability as key prey item.

This study evaluated the oxidative stress response in two cold-water fish species, the Antarctic Harpagifer antarcticus and the sub-Antarctic H. bispinis, following exposure to single and combined stressors: polyvinyl chloride (PVC) microplastics and thermal increase. Fish were exposed for 24 h to two temperature regimes (2 °C and 5 °C for H. antarcticus; 8 °C and 12 °C for H. bispinis) and were orally administered a PVC microplastic solution (200 mg/L). Oxidative stress was assessed through transcriptional and enzymatic activity analyses of key antioxidant markers: catalase, glutathione peroxidase, superoxide dismutase, and glutathione reductase. In H. antarcticus, gill antioxidant gene expression decreased in response to both stressors when applied individually, while H. bispinis exhibited transcriptional upregulation under the same conditions. In H. antarcticus, enzymatic activity in gill tissues increased for all four enzymes following exposure to both microplastics and elevated temperature. In contrast, H. bispinis showed a differential enzymatic response: thermal stress induced CAT activity, whereas exposure to microplastics specifically increased GR activity. At the hepatic level, H. antarcticus displayed increased transcription of antioxidant genes following exposure to both stressors. In H. bispinis, transcriptional upregulation was limited to GR and SOD in response to microplastics. However, under the combined exposure of multiple stressors, an inactivation of the antioxidant enzyme response was observed in the gills. This could indicate a functional collapse of the antioxidant system under conditions of exacerbated acute stress. The observed responses raise concerns about the potential vulnerability of polar and subpolar fishes, considering their ecological importance in trophic networks and the increasing pressure from multiple anthropogenic stressors in a rapidly changing climate.

Rising ocean temperatures due to climate change, combined with the intensification of anthropogenic activity, may lead to changes in the physiology and distribution of native species. Compounding climate stress, microplastic particles (MPs) enter the oceans through wastewater and the breakdown of macroplastics. Depending on their composition, they can be harmful and act as a vehicle for toxic substances, although their effects on native Antarctic and subantarctic species are unknown. Notothenioid fish are members of this group and are found inside and outside Antarctica, such as the Harpagifer, which has adapted to the cold and is particularly sensitive to thermal increases. Here, we aimed to evaluate the innate immune response in the head kidney, spleen, and foregut of two notothenoid fish, Harpagifer antarcticus and Harpagifer bispinis, exposed to elevated temperatures and PVC (polyvinyl chloride) microplastics. Adults from both species were collected on King George Island (Antarctica) and Punta Arenas (Chile), respectively. Specimens were assigned to a control group or exposed to a temperature increase (TI) or PVC microplastics (MPs), separately or in combination (MPs + TI). MP exposures were oral (gavage) for 24 h or aqueous (in a bath) for 24 and 48 h. Using real-time qPCR, we evaluated the relative gene expression of markers involved in the innate immune response, including tlr2 (toll-like receptor 2), tlr4 (toll-like receptor 4), myd88 (myeloid differentiation factor 88), nfkb (nuclear factor kb), il6 (interleukin 6), and il8 (irterleukin 8). We found differences between treatments when H. antarcticus and H. bispinis were exposed independently to MPs or thermal increase (TI) in the experiment with a cannula, showing an up-regulation in transcripts. In contrast, a down-regulation was observed when exposed in combination to MP + TI, which looked to be tissue-dependent. However, transcripts related to innate immunity in the bath experiment increased when exposure to both stressors was combined, mostly at 48 h. These results highlight the importance of evaluating the effects of multiple stressors, both independently and in combination, and whether these species will have the capacity to adapt or survive under these conditions, especially in waters where temperature is increasing and pollution is also rising, primarily from MP-PVC, a plastic widely used in various industries and among the population.

Marine heatwaves (MHWs) pose severe threats to coastal ecosystems. Fiddler crabs, key intertidal species in mangrove environments, are particularly vulnerable to MHWs in disturbed habitats, where vegetation degradation amplifies environmental stressors such as temperature and salinity. We conducted a biomarker-based assessment of the behavioral, physiological, and biochemical responses of fiddler crab species to MHWs in undisturbed and disturbed mangrove ecosystems. We also used a laboratory-based simulation of MHWs to investigate how environmental stressors (elevated temperature, salinity, and hypoxia) trigger behavioral responses of species in degraded habitats. Our findings reveal water escape and aggregation behaviors in crabs inhabiting disturbed mangroves during maximum high tide (MHW) events. These responses were triggered when burrow temperatures approached crabs' critical thermal maximum, suggesting a thermally induced escape mechanism. Crabs aggregating above the water level exhibited significant osmotic disruption, including a pronounced loss of osmoregulatory capacity, which heightened their vulnerability to desiccation. These physiological impairments were accompanied by signs of neuromuscular dysfunction, as indicated by altered acetylcholinesterase activity. In contrast, crabs from undisturbed mangrove habitats did not exhibit such behavioral or physiological disruptions. The laboratory experiments further demonstrated that the likelihood of escape and aggregation behaviors increased at lower temperatures under high-salinity and low-oxygen-concentration conditions. These results suggest that undisturbed mangrove habitats act as thermal refuges, mitigating the extreme effects of MHWs on the intertidal fauna.

Thermal variations due to global climate change are expected to modify the distributions of marine ectotherms, with potential pathogen translocations. This is of particular concern at high latitudes where cold-adapted stenothermal fish such as the Notothenioids occur. However, little is known about the combined effects of thermal fluctuations and immune challenges on the balance between cell damage and repair processes in these fish. The aim of this study was to determine the effect of thermal variation on specific genes involved in the ubiquitination and apoptosis pathways in two congeneric Notothenioid species, subjected to simulated bacterial and viral infections. Adult fish of Harpagifer bispinis and Harpagifer antarcticus were collected from Punta Arenas (Chile) and King George Island (Antarctica), respectively, and distributed as follows: injected with PBS (control), LPS (2.5 mg/kg) or Poly I:C (2 mg/kg) and then submitted to 2, 5 and 8 °C. After 1 week, samples of gills, liver and spleen were taken to evaluate the expression by real-time PCR of specific genes involved in ubiquitination (E3-ligase enzyme) and apoptosis (BAX and SMAC/DIABLO). Gene expression was tissue-dependent and increased with increasing temperature in the gills and liver while showing an opposite pattern in the spleen. Studying a pair of sister species that occur across the Antarctic Polar Front can help us understand the particular pressures of intertidal lifestyles and the effect of temperature in combination with biological stressors on cell damage and repair capacity in a changing environment.

Upwelling phenomena alter the physical and chemical parameters of the sea's subsurface waters, producing low levels of temperature, pH and dissolved oxygen, which can seriously impact the early developmental stages of marine organisms. To understand how upwelling can affect the encapsulated development of the gastropod Acanthina monodon, capsules containing embryos at different stages of development (initial, intermediate and advanced) were exposed to upwelling conditions (pH = 7.6; O2 = 3 mg L-1; T degrees = 9 degrees C) for a period of 7 days. Effects of treatment were determined by estimating parameters such as time to hatching, number of hatchlings per capsule, percentage of individuals with incomplete development, and shell parameters such as shell shape and size, shell strength, and the percentage of the organic/inorganic content.We found no significant impacts on hatching time, number of hatchlings per capsule, or percentage of incomplete development in either the presence or absence of upwelling, regardless of developmental stage. On the other hand, latent effects on encapsulated stages of A. monodon were detected in embryos that had been exposed to upwelling stress in the initial embryonic stage. The juveniles from this treatment hatched at smaller sizes and with higher organic content in their shells, resulting in a higher resistance to cracking 30 days after hatching, due to greater elasticity.Geometric morphometric analysis showed that exposure to upwelling condition induced a change in the morphology of shell growth in all post-hatching juveniles (0-30 days), regardless of embryonic developmental stage at the time of exposure. Thus, more elongated shells (siphonal canal and posterior region) and more globular shells were observed in newly hatched juveniles that had been exposed to the upwelling condition. The neutral or even positive upwelling exposure results suggests that exposure to upwelling events during the encapsulated embryonic phase of A. monodon development might not have major impacts on the future juvenile stages. However, this should be taken with caution in consideration of the increased frequency and intensity of upwelling events predicted for the coming decades.

Anchialine systems are coastal groundwater habitats around the world which host a unique community of cave adapted species (stygobionts). Such communities are expected to be separated by haloclines into either fresh or saline groundwater communities, hence climate changes (e.g., eustatic sea level shifts) and anthropic driven changes (e.g., salinization) may have a great impact on these stygobiont communities. Here we used cave-restricted species of Typhlatya from the Yucatan Peninsula as models to identify physiological capacities that enable the different species to thrive in marine groundwater (T. dzilamensis) or fresh groundwater (T. mitchelli and T. pearsei), and test if their distribution is limited by their salinity tolerance capacity. We used behavior, metabolic rates, indicators of the antioxidant system and cellular damage, and lactate content to evaluate the response of individuals to acute changes in salinity, as a recreation of crossing a halocline in the anchialine systems of the Yucatan Peninsula. Our results show that despite being sister species, some are restricted to the freshwater portion of the groundwater, while others appear to be euryhaline.

Marine heatwaves (MHW) pose an increasing threat and have a critical impact on meroplanktonic organisms, because their larvae are highly sensitive to environmental stress and key for species' dispersion and population connectivity. This study assesses the effects of MHW on two key moulting cycle periods within first zoea of the valuable crab, Metacarcinus edwardsii. First, the changes in swimming behaviour during zoea I were recorded and associated to moult cycle substages. Then, larvae were exposed during the zoea I to (1) control temperature of 12 degrees C, (2) Early MHW, occurring in intermoult, (3) Late MHW, occurring in premoult and (4) 14 degrees C, representing MHW during whole development. Additionally, optimum temperature was estimated from thermal performance curves through swimming behaviour of one-day zoea I. The timing of the MHW within the moulting cycle significantly affects larval fitness. Early MHW led to improved survival rates (72%) and reduced developmental times (9.8 days) compared to those exposed to Later MHW (63% and 10.3 days, respectively). As optimum temperature was higher than 12 degrees C, MHW events maybe favouring larval performance. These results highlight the importance of interaction between the moult cycle and environmental variables as a factor of sublethal effects on population dynamics.

Human settlements within the Antarctic continent have caused significant coastal pollution by littering plastic. The present study assessed the potential presence of microplastics in the gastrointestinal tract of the Antarctic fish Harpagifer antarcticus, endemic to the polar region, and in the sub-Antarctic fish Harpagifer bispinis. H. antarcticus. A total of 358 microfibers of multiple colors were found in 89 % of H. antarcticus and 73 % of H. bispinis gastrointestinal track. A Micro-FTIR analysis characterized a sub-group (n = 42) of microfibers. It revealed that most of the fibers were cellulose (69 %). Manmade fibers such as microplastics polyethylene terephtalate, acrylics, and semisynthetic/natural cellulosic fibers were present in the fish samples. All the microfibers extracted were textile fibers of blue, black, red, green, and violet color. Our results suggest that laundry greywater discharges of human settlements near coastal waters in Antarctica are a major source of these pollutants in the Antarctic fish.

Dissolved oxygen (DO) and water temperature vary in coastal environments. In tropical regions, the ability of aquatic ectotherms to cope with hypoxia and high-temperature interactive effects is fundamental for their survival. The mechanisms underlying both hypoxia and thermal tolerance are known to be interconnected, therefore, the idea of cross-tolerance between both environmental stressors has been put forward. We investigated the combined role of hypoxia and temperature changes on the physiological responses of blue crab Callinectes sapidus living in the southern Gulf of Mexico. We measured oxygen consumption, plasmatic biochemical indicators, total hemocyte count (THC), and antioxidant activity biomarkers in muscle and gill tissues of blue crab acclimated to moderate hypoxia or normoxia and exposed to a thermal fluctuation or a constant temperature, the former including a temperature beyond the optimum range. Animals recovered their routine metabolic rate (RMR) after experiencing thermal stress in normoxia, reflecting physiological plasticity to temperature changes. In hypoxia, the effect of increasing temperature was modulated as reflected in the RMR and plasmatic biochemical indicators concentration, and the THC did not suggest significant alterations in the health status. In both DO, the antioxidant defense system was active against oxidative (OX) damage to lipids and proteins. However, hypoxia was associated with an increase in the amelioration of OX damage. These results show that C. sapidus can modulate its thermal response in a stringent dependency with DO, supporting the idea of local acclimatization to tropical conditions, and providing insights into its potential as invasive species.

Sex-biased fishing mortality caused by male-only fishing may result in sperm limitation and recruitment overfishing. These effects can be observed by studying the size at functional reproduction (SFR) of lithodid females. We studied changes in the ratio of males to females, size truncation, and changes in SFR in the southern king crab (SKC) in two areas experiencing different fishing pressures in southern Chile. SFR was estimated with the Richards Function, a generalized 4-parameter logistic model. The shrinking of SFR of females may occur due to the proportional shrinking of reproductive and non-reproductive females and/or because the shape of the ogive changes, setting the inflection point back to smaller sizes. In the region with low fishing pressure the functional reproductive ogive was symmetric (gamma = 1, and the asymptote was similar to 1, indicating a balance in male availability. In the region with high fishing pressure gamma was 82 % lower and the asymptote was 23 % lower, an indicator of sperm limitation. The symmetric shape of the logistic curve, which is widely assumed when fitting maturity ogives, is not the most appropriate assumption in SKC and possibly in other males-only crab fisheries. More general shapes of the ogive should be estimated and studied.

Mitochondrial respirometry is key to understanding how environmental factors model energetic cellular processes. Until now, no reports have shown temperature effects and other environmental variables on cephalopod mitochondria activity because of the lack of a method to evaluate mitochondrial respiratory parameters on those groups of species. In this sense and for the first time, it showed the mitochondrial respirometry data of adult Octopus maya’s heart. Following the protocol is illustrated a step-by-step procedure to get the corresponding respiratory parameters.

Mitochondrial respirometry is key to understand how environmental factors model energetic cellular process. In the case of ectotherms, thermal tolerance has been hypothesized to be intimately linked with mitochondria capability to produce enough adenosine triphosphate (ATP) to respond to the energetic demands of animals in high temperatures. In a recent study made in Octopus maya was proposed the hypothesis postulating that high temperatures could restrain female reproduction due to the limited capacity of the animals' heart to sustain oxygen flow to the body, affecting in this manner energy production in the rest of the organs, including the ovarium Meza-Buendia AK et al. (2021). Unfortunately, until now, no reports have shown temperature effects and other environmental variables on cephalopod mitochondria activity because of the lack of a method to evaluate mitochondrial respiratory parameters in those species' groups. In this sense and for the first time, this study developed a method to obtain mitochondrial respirometry data of adult Octopus maya's heart. This protocol illustrates a step-by-step procedure to get high yield and functional mitochondria of cephalopod heart and procedure for determining the corresponding respiratory parameters. The procedure described in this paper takes approximately 3 to 4 hours from isolation of intact mitochondria to measurement of mitochondrial oxygen consumption.

Dissolved oxygen (DO) and water temperature vary in coastal environments due to natural and anthropogenic drivers. In tropical regions, the ability of aquatic ectotherms to cope with low DO and high temperature interactive effects is fundamental for their survival. The mechanisms underlying both hypoxia and thermal tolerance are interconnected according to the oxygen-and capacity-limited tolerance (OCLLT) hypothesis. Therefore, the idea of cross-tolerance between both environmental stressors has been put forward. This study investigates the combined role of hypoxia and temperature changes on the physiological response of the blue crab Callinectes sapidus living in the southern Gulf of Mexico. Measurements were performed in oxygen consumption, hemolymph metabolite profile, total hemocyte count (THC), and antioxidant activity biomarkers in muscle and gill tissues of normoxia- and hypoxia-acclimated blue crabs exposed to a fluctuating or constant thermal regime, the former including temperature beyond the optimum range. Animals were able to recover their routine metabolic rate (RMR) after experiencing thermal stress in normoxia, reflecting physiological plasticity to temperature. Moreover, low DO induced metabolic suppression as a possible strategy to conserve energy and extend survival under environmental stress as reflected in the RMR and hemolymph metabolite concentration. Hypoxia did not affect THC, suggesting that health status was not altered. In both DO levels, the antioxidant defense system (ADS) was active against oxidative (OX) damage on lipids and proteins. However, hypoxia was associated with an increase in ADS biomarker concentrations in muscles and gills and oxidative (OX) amelioration damage. These results show that C. sapidus can modulate its thermal response in a stringent dependency with DO, supporting the idea of local adaptation to the environmental conditions typical of their ecological niche in the tropics and providing insights into its potential as invasive species.