The vast majority of coastal organisms have been facing multigenerational scenarios of fluctuatingly high pCO2 and Cd pollution in their natural habitats. However, the adaptive capacity of these organisms to such combined stressors and the underlying mechanisms remain poorly understood. In this study, we conducted a multigenerational experiment (F1-F4) to investigate the adaptive responses of the marine copepod Tigriopus japonicus to combined fluctuatingly high pCO2 and Cd exposure, along with the associated mechanisms. Our findings revealed that steady high pCO2 aggravated Cd multigenerational toxicity, and it was more under fluctuating acidification. Notably, by the F4 generation, copepods potentially adapted to the combined stressors. Through transcriptomic and DNA methylation analyses of copepods from the F1 and F4 generations, we found that under combined exposure, F1 copepods likely reallocated more energy to counteract Cd toxicity; however, DNA hypermethylation inhibited Cd exclusion and detoxification/stress response pathways, ultimately compromising development and reproduction. In contrast, in the F4 generation, DNA hypomethylation enhanced processes such as cuticle repair program, compensatory mechanism (e.g., detoxification and immune response), and reproduction, consequently increasing the copepod's fitness. These findings reveal an epigenetic basis for phenotypic acclimatization, offering marine copepods a supplementary mechanism to cope with combined stressors.

Despite the great interest in the consequences of global change stressors on marine organisms, their interactive effects on cadmium (Cd) bioaccumulation/biotoxicity are very poorly explored, particularly in combination with the toxicokinetic model and molecular mechanism. According to the projections for 2100, this study investigated the impact of elevated pCO2 and increased temperature (isolated or joint) on Cd uptake dynamics and transcriptomic response in the marine copepod Tigriopus japonicus. Toxicokinetic results showed significantly higher Cd uptake in copepods under increased temperature and its combination with elevated pCO2 relative to the ambient condition, linking to enhanced Cd bioaccumulation. Transcriptome analysis revealed that, under increased temperature and its combination with elevated pCO2, up-regulated expression of Cd uptake-related genes but down-regulation of Cd exclusion-related genes might cause increased cellular Cd level, which not only activated detoxification and stress response but also induced oxidative stress and concomitant apoptosis, demonstrating aggravated Cd biotoxicity. However, these were less pronouncedly affected by elevated pCO2 exposure. Therefore, temperature seems to be a primary factor in increasing Cd accumulation and its toxicity in the future ocean. Our findings suggest that we should refocus the interactive effects between climate change stressors and Cd pollution, especially considering temperature as a dominant driver.

Marine dinoflagellates are increasingly affected by ongoing global climate changes. While understanding of their physiological and molecular responses to individual stressors anticipated in the future ocean has improved, their responses to multiple concurrent stressors remain poorly understood. Here, we investigated the individual and combined effects of elevated temperature (26 degrees C relative to 22 degrees C), increased p CO 2 (1000 mu atm relative to 400 mu atm), and high nitrogen: phosphorus ratio (180:1 relative to 40:1) on a harmful algal bloom-causing dinoflagellate Prorocentrum obtusidens under short-term (28 days) exposure. Elevated temperature was the most dominant stressor affecting P. obtusidens at physiological and transcriptomic levels. It significantly increased cell growth rate and maximum photosynthetic efficiency (Fv/Fm), but reduced chlorophyll a , particulate organic carbon, particulate organic nitrogen, and particulate organic phosphorus. Elevated temperature also interacted with other stressors to produce synergistic positive effects on cell growth and Fv/Fm. Transcriptomic analysis indicated that elevated temperature promoted energy production by enhancing glycolysis, tricarboxylic acid cycle, and nitrogen and carbon assimilation, which supported rapid cell growth but reduced material storage. Increased p CO 2 enhanced the expression of genes involved in ionic acid-base regulation and oxidative stress resistance, whereas a high N:P ratio inhibited photosynthesis, compromising cell viability, although the effect was alleviated by elevated temperature. The combined effect of these multiple stressors resulted in increased energy metabolism and up-regulation of material-synthesis pathways compared to the effect caused by elevated temperature alone. Our results underscore ocean warming as the predominant stressor for dinoflagellates and highlight the complex, synergistic effects of multi-stressors on dinoflagellates.

Paralytic shellfish toxins (PSTs) can pose a serious threat to human health. Among them, saxitoxin (STX) is one of the most potent natural neurotoxins. Here, the copepod Tigriopus japonicus, was exposed to environmentally relevant concentrations (2.5 and 25 μg/L) STX for 48 h. Although no lethal effects were observed at both concentrations, the transcriptome was significantly altered, and displayed a concentration-dependent response. STX exposure decreased the copepod's metabolism and compromised immune defense and detoxification. Additionally, STX disturbed signal transduction, which might affect other cellular processes. STX exposure could inhibit the copepod's chitin metabolism, disrupting its molting process. Also, the processes related to damage repair and protection were up-regulated to fight against high concentration exposure. Collectively, this study has provided an early warning of PSTs for coastal ecosystem not only because of their potent toxicity effect but also their bioaccumulation that can transfer up the food chain after ingestion by copepods.

Marine ecosystem has been experiencing multiple stressors caused by anthropogenic activities, including ocean acidification (OA) and nickel (Ni) pollution. Here, we examined the individual/combined effects of OA (pCO2 1000 μatm) and Ni (6 μg/L) exposure on a marine copepod Tigriopus japonicus for six generations (F1-F6), followed by one-generation recovery (F7) in clean seawater. Ni accumulation and several important phenotypic traits were measured in each generation. To explore within-generation response and transgenerational plasticity, we analyzed the transcriptome profile for the copepods of F6 and F7. The results showed that Ni exposure compromised the development, reproduction and survival of copepods during F1-F6, but its toxicity effects were alleviated by OA. Thus, under OA and Ni combined exposure, due to their antagonistic interaction, the disruption of Ca2+ homeostasis, and the inhibition of calcium signaling pathway and oxytocin signaling pathway were not found. However, as a cost of acclimatization/adaption potential to long-term OA and Ni combined exposure, there was a loss of transcriptome plasticity during recovery, which limited the resilience of copepods to previously begin environments. Overall, our work fosters a comprehensive understanding of within- and transgenerational effects of climatic stressor and metal pollution on marine biota.

The continuous fragmentation of plastics and release of synthetic nanoplastics from products have been aggravating nanoplastic pollution in the marine ecosystem. The carrier role of nanoplastics may increase the bioavailability and toxicity effects of toxic metals, e.g., mercury (Hg), which is of growing concern. Here, the copepod Tigriopus japonicus was exposed to polystyrene nanoplastics (PS NPs) and Hg (alone or combined) at environmental realistic concentrations for three generations (F0-F2). Then, Hg accumulation, physiological endpoints, and transcriptome were analyzed. The results showed that the copepod's reproduction was significantly inhibited under PS NPs or Hg exposure. The presence of PS NPs caused significantly higher Hg accumulation, lower survival, and lower offspring production in copepods relative to Hg exposure, suggesting an increased threat to the copepod's survivorship and health. From the molecular perspective, combined PS NPs and Hg caused a graver effect on the DNA replication, cell cycle, and reproduction pathways relative to Hg exposure, linking to lower levels of survivorship and reproduction. Taken together, this study provides an early warning of nanoplastic pollution for the marine ecosystem not only because of their adverse effect per se but also their carrier role for increasing Hg bioaccumulation and toxicity in copepods.

Coastal waters have experienced fluctuations in partial pressure of carbon dioxide (pCO2) and mercury (Hg) pollution, yet little is known concerning how natural pCO2 fluctuations affect Hg biotoxicity. Here, a marine copepod Tigriopus japonicus was interactively exposed to different seawater pCO2 (ambient 400, steady elevated 1000, and fluctuating elevated 1000 ± 600 μatm) scenarios and Hg (control, 2 μg/L) treatments for 7 d. The results showed that elevated pCO2 decreased Hg bioaccumulation, and it was even more under fluctuating elevated pCO2 condition. We found energy depletion and oxidative stress under Hg-treated copepods, while combined exposure initiated compensatory response to alleviate Hg toxicity. Intriguingly, fluctuating acidification presented more immune defense related genes/processes in Hg-treated copepods when compared to steady acidification, probably linking with the greater decrease in Hg bioaccumulation. Collectively, understanding how fluctuating acidification interacts with Hg contaminant will become more crucial in predicting their risks to coastal biota and ecosystems.

Growing evidence demonstrates that global change can modulate mercury (Hg) toxicity in marine organisms; however, the consensus on such effect is lacking. Here, we conducted a meta-analysis to evaluate the effects of global change stressors on Hg biotoxicity according to the IPCC projections (RCP 8.5) for 2100, including ocean acidification (-0.4 units), warming (+4 °C), and their combination (acidification-warming). The results indicated an overall aggravating effect (ln RRΔ = -0.219) of global change on Hg toxicity in marine organisms, while the effect varied with different stressors; namely, acidification potentially alleviates Hg biotoxicity (ln RRΔ = 0.117) while warming and acidification-warming have an aggravating effect (ln RRΔ = -0.328 and -0.097, respectively). Moreover, warming increases Hg toxicity in different trophic levels, i.e., primary producers (ln RRΔ = -0.198) < herbivores (ln RRΔ = -0.320) < carnivores (ln RRΔ = -0.379), implying increasing trends of Hg biomagnification through the food web. Notably, ocean hypoxia appears to boost Hg biotoxicity, although it was not considered in our meta-analysis because of the small sample size. Given the persistent global change and combined effects of these stressors in marine environments, multigeneration and multistressor research is urgently needed to fully disclose the impacts of global change on Hg pollution and its risk.

Plastic pollution is a serious problem in the global marine environment because it can produce negative effects at the biological and ecological levels. Due to large surface-area-to-volume ratio and inherent hydrophobicity, nanoplastics can serve as carriers of contaminants, and may affect their fate and toxicity in marine environments. However, the combined effects of nanoplastics and mercury (Hg) in marine organisms have not been well characterized. In this study, after verifying the ingestion of polystyrene nano-size plastics (PS NPs, 50 nm) by the copepod Tigriopus japonicus and adsorption of Hg to PS NPs, we investigated the effects of PS NPs and Hg exposure (alone or in combination) for 48 h on the copepods. Specifically, a 72-h depuration was performed after 48 h exposure. The results showed that after 48 h exposure, the copepod's Hg concentration was significantly increased in the combined exposure group compared to that in the Hg treatment group, but these differences did not persist following 24 h of depuration. Therefore, PS NPs transported Hg into the copepods but did not promote Hg bioaccumulation. Treatment with PS NPs alone did not induce toxicity in T. japonicus, but co-exposure to PS NPs and Hg resulted in elevated transcription of genes related to energy production, antioxidant response, and detoxification/stress defense when compared with Hg treatment alone, demonstrating the synergistic interaction between PS NPs and Hg. Our findings contribute to a comprehensive understanding about the combined toxicity of nanoplastics and metals and the potential ecological risks of associated with these effects in marine environments.

Coastal systems experience diel fluctuation of pCO2 and cadmium (Cd) pollution; nevertheless, the effect of fluctuating pCO2 on Cd biotoxicity is poorly known. In this study, we initially performed the isotopically enriched organism bioassay to label Tigriopus japonicus with 113Cd (5 μg/L) to determine the Cd accumulation rate constant (kaccu) under ambient (400 μatm) and steadily (1000 μatm) and fluctuatingly elevated (1000 ± 600 μatm) pCO2 conditions for 48 h. Next, T. japonicus was interactively subjected to the above pCO2 exposures at Cd (control, 5, and 500 μg/L) treatments for 7 d. Biochemical and physiological responses for copepods were analyzed. The results showed that steadily increased pCO2 facilitated Cd bioaccumulation compared to ambient pCO2, and it was more under fluctuating acidification conditions. Despite compensatory reactions (e.g., increased energy production), Cd ultimately induced oxidative damage and apoptosis. Meanwhile, combined treatment exhibited higher toxicity (e.g., increased apoptosis) relative to Cd exposure, and even more if fluctuating acidification was considered. Intriguingly, fluctuating acidification inhibited Cd exclusion in Cd-treated copepods compared to steady acidification, linking to higher Cd kaccu and bioaccumulation. Collectively, CO2-driven acidification could aggravate Cd toxicity, providing a mechanistic understanding of the interaction between seawater acidification and Cd pollution in marine copepods.

Here, we examined the 48-h acute toxicity of cadmium (Cd) in the marine copepod Tigriopus japonicus under two pCO2 concentrations (400 and 1000 μatm). Subsequently, T. japonicus was interactively exposed to different pCO2 (400, 1000 μatm) and Cd (control, 500 μg/L) treatments for 48 h. After exposure, biochemical and physiological responses were analyzed for the copepods. The results showed that the 48-h LC50 values of Cd were calculated as 12.03 mg/L and 9.08 mg/L in T. japonicus, respectively, under 400 and 1000 μatm pCO2 conditions. Cd exposure significantly promoted Cd exclusion/glycolysis, detoxification/stress response, and oxidative stress/apoptosis while it depressed that of antioxidant capacity. Intriguingly, CO2-driven acidification enhanced Cd bioaccumulation and its toxicity in T. japonicus. Overall, our study provides a mechanistic understanding about the interaction between seawater acidification and Cd pollution in marine copepods.