Divalent metal transporter 1 (DMT1) is thought to be the primary route for non-heme iron absorption in vertebrates, but its systemic role remains poorly understood. Using CRISPR-Cas9 gene editing, we generated a DMT1 knockout (dmt1-/-; KO) zebrafish mutant line to examine the developmental and physiological consequences of DMT1 loss. Phenotypic and hematological assessments were performed alongside measurements of whole-body and tissue-specific metal concentrations. Further, to identify potential compensatory pathways during DMT1 loss, the expression profile of candidate metal transporters or ion channels (hcp1, zip4, zip8, zip14, and ecac) was quantified using droplet digital PCR (ddPCR). DMT1 KO larvae exhibited delayed development, anemia, and broad disruption in multiple trace metals (iron, zinc, manganese, cobalt, and selenium). Gene expression analysis during early development revealed higher hcp1 mRNA abundance in the mutant, suggesting a possible compensatory response to maintain metal homeostasis during DMT1 loss. Although viable to adulthood, the mutants had persisting iron dysregulation and red blood cell abnormalities. This study provides the first in vivo evidence of the physiological role of DMT1 in multi-metal balance in fish and offers new insight into compensatory mechanisms underlying DMT1 deficiency.
Since the 1960s, global plastic production has increased dramatically, resulting in widespread accumulation and fragmentation of plastic debris in terrestrial, aquatic, and atmospheric systems. Microplastics (MPs), originating from primary sources such as industrial raw materials and microbeads, as well as secondary sources from fragmentation of larger plastics, have emerged as contaminants of particular concern due to their persistence, high abundance, and capacity to interact with diverse organisms. This review provides a comprehensive overview of microplastic behavior, focusing on fish as key bioindicators. We summarize the environmental distribution of microplastics, their uptake pathways, bioaccumulation and potential trophic transfer, organ-level and cellular translocation, and clearance mechanisms in fish, integrating both laboratory and field observations. Advanced analytical and imaging techniques-including fluorescence labeling, molecular and metal probes, isotopic tracing, hyperspectral imaging, and surface-enhanced Raman spectroscopy-are evaluated for their advantages, limitations, and reliability in detecting MPs. Furthermore, we synthesize evidence on the multifaceted effects of MPs on fish growth, development, behavior, antioxidant capacity, immune function, and gut microbiota. This review uniquely highlights the full pathway of MPs from environmental exposure to organismal impact, providing critical insights into their ecological risks, methodological considerations, and implications for environmental monitoring and pollution mitigation.
Metamifop (MET) is frequently detected in aquatic environments and threatens non-target aquatic organisms, yet chronic toxicological data on aquatic invertebrates remain scarce. To evaluate the physiological and molecular mechanisms of MET toxicity, Daphnia magna was subjected to 21-day chronic exposure at 0, 0.4, 4, and 40 μg/L. Results showed a pronounced concentration-dependent growth and reproductive toxicity, evidenced by a reduced the offspring number at the first brood in the 0.4 μg/L treatment, while 4 and 40 μg/L MET further decreased the total number of offspring per female and the body length of D. magna. MET impaired digestive function by altering the mRNA expression levels of α-amylase, trypsin, and α-esterase, disrupted energy metabolism by downregulating ak expression, lowering ATP content, and decreasing ATPase activity, and inhibited metabolic detoxification mechanisms via reduced transcriptional levels of cyp360a8, gst, and p-gp genes. Exposure to 40 μg/L MET also significantly decreased the swimming speed of D. magna. MET further induced oxidative stress in D. magna, as supported by diminished SOD and CAT activity, downregulated transcription of trx and trxr, decreased levels of hsp70 and hsp90, and elevated contents of GSH and MDA. Reduced expression of the nvd, ecr, cyp314, and vtg genes indicated impairment of ecdysteroid signaling system. Collectively, these findings demonstrate that prolonged MET exposure exerts adverse effects on growth and reproduction that are closely associated with disturbed energy metabolism, detoxifying capacity, antioxidant defense, and ecdysteroid signaling system in D. magna, highlighting the ecological hazards of MET to aquatic ecosystems.
Microplastic pollution, particularly from polyethylene (PE), poses an increasing threat to coastal ecosystems, yet how particle size and exposure duration jointly regulate organismal responses remains poorly understood. Here, we investigated the size- and time-dependent toxic effects of PE-MPs (10 μm and 50 μm) on the Kumamoto oyster (Crassostrea sikamea) using an integrative framework combining physiological biomarkers (SOD, CAT, MDA), histopathology (gills and hepatopancreas), transcriptomics (gills), and metabolomics (hepatopancreas) during acute (1 day), short-term (7 days), and long-term (14 days) phases. Both PE-MP sizes induced significant oxidative stress and tissue injury in a time-dependent manner, with smaller particles casing more persistent oxidative stress, greater metabolic disturbance, and stronger immune suppression. Multi-omics analyses revealed a clear phase-dependent response pattern characterized by early defense activation, short-term metabolic reprogramming, and long-term functional suppression. Acute exposure activated oxidative stress responses, cytoskeletal remodeling, and particle clearance-related pathways, whereas short-term exposure was associated with metabolic reprogramming characterized by enhanced glycolysis and amino acid metabolism, suggesting increased energetic demands during stress responses. In contrast, long-term exposure resulted in coordinated suppression of immune, digestive, and lipid metabolic-related pathways, together with a metabolic shift toward long-term energy conservation. Overall, these findings suggest that PE-MPs exposure may induce coordinated physiological and metabolic adjustments associated with energy trade-offs under chronic stress conditions, highlighting the importance of particle size and exposure duration in ecological risk assessment for coastal and aquaculture environments.
Esketamine hydrochloride is increasingly used as a rapid-acting antidepressant, and its expanding clinical and non-medical use has raised concerns regarding its release into aquatic systems via wastewater treatment plant effluents as an emerging psychoactive contaminant. However, its potential neurodevelopmental toxicity in aquatic organisms remains insufficiently characterized. In this study, zebrafish embryos were exposed to esketamine hydrochloride during early development, and its toxic effects were evaluated using an integrated framework combining developmental, behavioral, histological, transcriptomic, oxidative stress-related, and apoptosis-related endpoints. Early-life esketamine exposure altered multiple developmental indicators, including head length, eye depth, interocular distance, and body length, and disrupted locomotor regulation at later stages, particularly light-dark responsiveness and spatial preference. Histological examination further revealed exposure-related alterations in brain tissue organization. Transcriptomic profiling identified coordinated changes in pathways associated with redox homeostasis, protein synthesis, and phototransduction-related signaling. Targeted validation demonstrated significant upregulation of oxidative stress-related genes, including sod1 and sod2, while ELISA-based assays showed exposure-dependent alterations in SOD, CAT, GSH, and MDA levels. Acridine orange (AO) staining showed increased apoptosis-related fluorescence signals in the head region, with AO-positive puncta density differing significantly among groups. Integrative correlation analysis further linked developmental, behavioral, oxidative stress-related, and apoptosis-related endpoints. Notably, these effects occurred in the absence of overt lethality. Collectively, these findings demonstrate that esketamine interferes with neurodevelopmental and behavioral processes in zebrafish larvae and support the incorporation of early-life neurobehavioral endpoints into risk assessment frameworks for neuroactive pharmaceuticals.
Organic UV filters are ubiquitous aquatic contaminants, yet their sublethal, age-specific effects on non-target invertebrates remain poorly understood. This study investigated the ontogenetic shifts in detoxification strategies of Daphnia magna exposed to three common UV filters (Avobenzone, Octocrylene, Octinoxate) and their environmentally relevant ternary mixture. By comparing 3-day-old juveniles and 6-day-old subadults, we integrated molecular biomarkers (in vivo ECOD and MXR efflux activity, and CAT/SOD antioxidant enzymes) with whole-organism physiological endpoints (heart rate and locomotion). Results revealed a distinct developmental window of susceptibility. Juveniles exhibited a highly responsive phenotype, characterized by robust MXR capacity and highly inducible CYP450 enzymes and active antioxidant mobilization. However, this active biochemical state was accompanied by pronounced bradycardia under chemical stress, consistent with the physiological cost of detoxification. In contrast, 6-day-old sub-adults displayed ttenuated molecular response pattern, showing lower biochemical inducibility but maintaining stable cardiac function. Alarmingly, Avobenzone acted as a dual-action toxicant in adults, simultaneously inhibiting both CYP metabolism and MXR efflux. Furthermore, the low-dose mixture compromised the adult MXR system without triggering explicit avoidance behaviour. These findings highlight that UV filters act as active modulators of the Daphnia defence network, emphasizing the need to incorporate life-stage specific vulnerabilities and mechanistic biomarkers into future ecological risk assessments.
To identify the differential properties of anti-microbial peptides (AMPs) and proteins in the burying beetle Nicrophorus quadripunctatus and the short sexton beetle Necrodes littoralis, we performed comparative transcriptomic analyses, quantified gene expression levels, examined structural variations, and evaluated anti-microbial and hemolytic activities of representative AMPs, including cecropin B and defensin. Two AMPs (cecropin B and defensin) and four anti-microbial proteins (cathelicidin, lysozyme, major royal jelly protein, and peptidoglycan recognition proteins) were predominantly detected. Defensin was the most highly expressed AMP in both species, whereas cecropin B showed relatively lower expression. Cecropin B and defensin in N. quadripunctatus exhibited 52.2- and 100.9-fold higher relative transcription levels, respectively, compared with N. littoralis, representing the two largest expression differences among the examined genes. Notably, substantial sequence variation was observed in the mature peptide regions of both AMPs between the two species. The α-helices of both AMPs from N. quadripunctatus exhibited longer segments than those from N. littoralis as a structural feature. Defensin of N. littoralis exhibited significantly higher anti-microbial activity against Gram-positive bacteria compared with that of N. quadripunctatus. In addition, only cecropin B exhibited anti-microbial activity against Gram-negative bacteria, and neither of the two AMPs showed detectable hemolytic activity. Overall, this study provides the first comparative transcriptomic, structural, and functional characterization of AMPs and anti-microbial proteins in N. quadripunctatus and N. littoralis, offering important foundational insights for the future development of bacteria-specific AMPs as alternatives to conventional antibiotics.
Microplastics have been shown to alter physiological processes in microalgae, key primary producers in aquatic systems. Although microalgae rarely exist in isolation, most studies evaluating microplastic toxicity have relied on monoculture systems, limiting insight into how interspecific interactions influence algal responses. In this study, we directly compared the physiological and morphological responses of two green microalgae, Chlamydomonas reinhardtii and Chlorella vulgaris, to high-density polyethylene (HDPE) microplastics (1000 mg/L; 20 μm) under mono- and co-culture conditions for six days. Algal responses were assessed using cell density, photosynthetic activity, and cell morphology as key endpoints to evaluate how interspecific interactions modulate species-specific sensitivity to microplastic exposure. HDPE microplastics consistently affected C. reinhardtii regardless of culture conditions, whereas alterations in C. vulgaris were more pronounced under co-culture conditions, indicating enhanced sensitivity under interspecific interactions. Overall, HDPE microplastics disrupted algal biomass and reshaped interspecific interactions under co-culture conditions, highlighting the importance of incorporating interspecific interactions into microplastic toxicity assessments of microalgae.
The widespread use of pharmaceuticals has led to their frequent detection in aquatic environments, raising concerns about adverse effects on non-target organisms. This study examined behavioral and endocrine responses to acute repetitive exposure (4 days, 15 min day-1) to environmentally relevant concentrations of azithromycin (AZT, 12.5 μg mL-1), nimesulide (NIME, 0.4 μg mL-1), and their combination in zebrafish (Danio rerio). Behavior was assessed using the Novel Tank Test, Social Preference Test, and Light/Dark Test. Whole-body cortisol, acetylcholinesterase (AChE) and catalase (CAT) activities, lipid peroxidation (TBARS), and non-protein thiols (NTP) were measured as stress biomarkers. NIME significantly reduced locomotion (∼30%) and time spent in upper zones (p < 0.05), indicating hypolocomotion and anxiety-like behavior. AZT elevated cortisol (p < 0.05) but caused minimal behavioral changes. Co-exposure intensified behavioral impairments and further increased cortisol (p < 0.01), suggesting synergistic effects. NIME increased TBARS, indicating oxidative damage. AChE activity was inhibited in the brain by NIME, while AZT and the combination increased systemic AChE. CAT activity rose in the combination group, possibly reflecting a compensatory response to reactive oxygen species. These findings demonstrate that short-term, repeated exposure to mixtures of commonly detected pharmaceuticals, even at environmentally relevant concentrations, can disrupt behavioral, biochemical, and endocrine homeostasis in aquatic organisms. This highlights the ecological risks posed by multi-compound contamination in aquatic environments.
The competing endogenous RNA (ceRNA) hypothesis provides a novel perspective for investigating the post-transcriptional regulatory mechanisms of detoxification enzyme genes. In this study, a cytochrome P450 gene, Rhopalosiphum padi CYP307A1 was significantly upregulated in the imidacloprid-resistant strain (RP-R) compared to the imidacloprid-susceptible strain (RP-S), and silencing CYP307A1 significantly increased the susceptibility of R. padi to imidacloprid. Both CYP307A1 and the long non-coding RNA lnc40545 containing conserved microRNA response elements (MREs) for miR-965 were predicted by bioinformatic analysis, and was experimentally validated by using a dual-luciferase reporter assay, showing that miR-965 specifically targets and binds to both CYP307A1 and lnc40545. Furthermore, silencing lnc40545 resulted in the significant downregulation of CYP307A1 and a concomitant increase in imidacloprid sensitivity in R. padi. These results indicate that lnc40545 acts as a “molecular sponge” to sequester miR-965, thereby indirectly upregulating the expression of CYP307A1, suggesting the involvement of a lncRNA-mediated ceRNA mechanism in imidacloprid resistance in R. padi. It not only advances our understanding of insect detoxification metabolism but also provides a crucial theoretical basis for the integrated management of imidacloprid resistance.
Trietazine is an s-triazine-based herbicide commonly applied in agricultural systems, but its stability in aquatic environments has prompted growing concern about unintended effects on non-target organisms. To assess its developmental toxicity and clarify associated mechanisms, we used both wild-type zebrafish embryos and multiple organ-specific transgenic lines as in vivo models of early vertebrate development. The embryos were continuously exposed to trietazine from 8 to 96 h post-fertilization, resulting in clear dose-dependent developmental defects, including shortened body length, reduced ocular size, and frequent pericardial and yolk sac edema, indicative of moderate to high embryotoxicity across the tested concentrations. In parallel, trietazine markedly enhanced intracellular reactive oxygen species levels and induced transcriptional activation of genes involved in inflammatory signaling and programmed cell death, supporting a central role for oxidative stress-driven apoptosis in its developmental toxicity. To delineate organ-level toxicity, we examined a panel of transgenic zebrafish lines. Embryos expressing cmlc2:dsRED and gata1a:dsRED exhibited pronounced cardiac malformations and impaired blood flow, while altered vascular organization was detected in the flk1:mCherry line. Disruption of cardiovascular development was further supported by aberrant expression patterns of the cardiac regulatory genes nkx2.5 and vmhc, as revealed by whole-mount in situ hybridization. Additionally, neurodevelopmental deficits and liver malformations were observed in olig2:dsRED, huC:EGFP, and fabp10a:dsRED embryos, demonstrating multi-organ developmental toxicity involving the nervous system and liver. Taken together, these results demonstrate that trietazine interferes with normal embryonic development across multiple organ systems by promoting oxidative stress-dependent inflammatory and apoptotic responses in both wild-type and transgenic zebrafish models. Our findings highlight the potential ecological hazards associated with trietazine contamination in aquatic ecosystems and provide mechanistic insight into the level and spectrum of its developmental toxicity.
Glyphosate is one of the most widely used herbicides globally and is frequently detected in surface waters in multiple regions. Despite its toxicological relevance and potential ecological and human health implications (e.g., carcinogenity), its use was recently renewed in the European Union until 2033. A growing body of literature has showed that glyphosate can cause adverse effects in mollusks, but previous studies have focused primarily on survival, reproduction, genotoxicity, and certain cellular biomarkers, leaving its effects on molluscan behavior and embryonic development largely unknown. In the present study, we investigated the developmental, behavioral, and biochemical consequences of chronic glyphosate exposure on embryos of the widely used molluscan model, the great pond snail (Lymnaea stagnalis). Embryos were exposed from the single-cell stage to hatching to environmentally relevant concentrations of glyphosate (25, 100, and 500 μg L-1). All applied concentrations transiently delayed hatching without causing morphological abnormalities. Moreover, the highest concentration transiently increased heart rate, locomotion (gliding), and feeding, indicating elevated metabolic requirements. Biochemical analyses in hatched embryos demonstrated that all exposure concentrations significantly reduced vitellogenin content and lipid peroxidation, whereas a significant increase in catalase and lactate dehydrogenase (LDH) activity was observed only at the highest concentration, suggesting altered energy allocation and oxidative stress. Histochemical staining confirmed increased LDH activity, revealing tissue-specific metabolic activity, while no histological alterations were observed in hatched embryos. Collectively, our findings reveal previously unrecognized sublethal effects of glyphosate on molluscan embryos (e.g., altered heart rate, behavior, and cellular biomarkers), highlighting that this compound can disrupt growth, energy metabolism, and cellular homeostasis in Lymnaea embryos with potential consequences for post-hatching performance, resilience, and long-term fitness.
Hepatic damage in fish induced by microplastic exposure has garnered increasing concern, yet its molecular mechanisms remain insufficiently elucidated. In this study, 30 days after hatching (dah) Nile tilapia were subjected to sub-chronic exposure to polystyrene microplastics (PS; 100 nm) for 14 days. Histopathological examination revealed evident inflammatory cell infiltration in the livers of PS-exposed fish compared to the control fish. Transmission electron microscopy showed elevated mitochondrial rupture and increased autophagosome formation. Immunofluorescence and Western blot analyses showed upregulated Lc3b and downregulated P62 protein levels, suggesting enhanced hepatic autophagy. Transcriptomic profiling of liver tissues and subsequent KEGG enrichment analysis highlighted significant upregulation of genes involved in the MAPK, NOD-like receptor, Toll-like receptor, and autophagy signaling pathways. Metabolomic profiling indicated notable enrichment in glutathione metabolism, ferroptosis, cysteine and methionine metabolism, and the NOD-like receptor pathway. Integrated transcriptomic and metabolomic KEGG analysis consistently identified ferroptosis as a centrally enriched pathway. Further gene expression and metabolite analyses demonstrated marked upregulation of immune-related genes, autophagy-related genes, and ferroptosis-pathway genes. Concurrently, ferroptosis-related metabolites including glutathione and cysteine were significantly decreased. Meanwhile, levels of lipid metabolites such as 2-oleoylglycerol were also reduced, whereas lipid peroxidation products represented by 4-hydroxynonenal were significantly increased. Additional validation confirmed increased expression of inflammatory factors (il-1β, tgf-β, nlrp3) and altered iron homeostasis in the PS-exposed fish liver. These findings indicate that sub-chronic PS exposure promotes hepatic ferroptosis via immune-mediated activation of autophagy, ultimately leading to liver injury in Nile tilapia. Our study provides novel insights into the mechanisms underlying microplastic-induced tissue damage in aquatic organisms.
Titanium dioxide nanoparticles (nano-TiO₂) and carbamazepine (CBZ) are both emerging contaminants of concern in coastal environments. Current monitoring data indicate that nano-TiO₂ occur in nearshore seawater affected by land-based inputs, and CBZ is persistently detected in estuarine and nearshore waters; therefore, these contaminants may co-occur at a regional scale. In this study, the thick-shelled mussel (Mytilus coruscus) was used as a model organism. Nominal concentrations of nano-TiO₂ (100 μg/L) and CBZ (10 μg/L) were applied in single and combined exposure experiments over 14 days. Particle characterization, gonadal morphology, sex steroid hormone levels, and key gene expression analyses were integrated to evaluate reproductive hormone-disrupting effects. The results showed that nano-TiO₂ readily aggregated in seawater, and distinct nano-TiO₂ aggregates were observed on the surface of CBZ crystals in dried mixed samples. Phenotypically, exposure to either pollutant alone reduced the gonadal area and gonadosomatic index, showing significant sex-specific differences in the CBZ group. In contrast, combined exposure caused more pronounced reproductive tissue damage. Hormonally, exposure disrupted sex steroid homeostasis, decreasing progesterone, estrone, and estradiol (E2), accompanied by increased testosterone (T). These hormonal changes exhibited sex-dependent patterns across treatments, with a decrease in E2 and an increase in T being more pronounced in the combined-exposure group. Molecularly, exposures significantly downregulated SF-1 and Wnt signaling genes (β-catenin, Wnt4), while upregulating Wnt7b. Overall, single exposure to nominal concentrations of nano-TiO₂ or CBZ was sufficient to induce sex-specific reproductive damage in M. coruscus, whereas combined exposure caused more pronounced changes at the morphological, hormonal, and transcriptional levels.
C-type lectins (CTLs), a pivotal family of PRRs, are essential for mediating host immune defense via specific recognition of exogenous pathogenic microbes. A new perlucin family member (designated SpPer), a typical member of the CTL superfamily, was cloned and characterized from the mud crab Scylla paramamosain. The complete coding region of SpPer was 582 bp in length, encoding a 193-amino acid protein that harbors a single carbohydrate-recognition domain (CRD), a signature structural feature of CTLs. Tissue-specific expression profiling confirmed prominent SpPer enrichment in the hepatopancreas and gill of healthy mud crabs. Quantitative real-time PCR assays demonstrated that SpPer mRNA transcription was induced in the hepatopancreas following Vibrio parahaemolyticus infection and ammonia nitrogen exposure, suggesting its potential involvement in immune responses and environmental stress adaptation. To elucidate the regulatory mechanism of SpPer, in vivo silencing was performed using RNA interference (RNAi). Following SpPer silencing, transcript abundances of several immune effector genes (anti-lipopolysaccharide factor, ALF; crustin antimicrobial peptide, crustin; Toll-like receptor, Toll; heat shock protein 70, HSP70; and superoxide dismutase, SOD) were significantly decreased, suggesting that SpPer played a vital role in immune defense by regulating genes involved in pathogen killing, signaling transduction, cellular protection, and antioxidant defense. Functional assays showed that SpPer silencing impaired bacterial clearance in the hemolymph and impaired the survival ability of mud crabs upon V. parahaemolyticus infection. Furthermore, under ammonia nitrogen stress, SpPer silencing led to a remarkable rise in cumulative mortality relative to the untreated control group. Taken together, these observations illustrate that SpPer is essential for regulating innate immune defense against bacterial infection and enhancing tolerance to ammonia stress in S.paramamosain.
Accumulating evidence indicates that exogenous toxins can induce lipid metabolic disorders by influencing the gut microbiome. However, the effects of ammonia nitrogen on the gut microbiota and hepatic lipid metabolism of the burbot (lota lota) remain unknown. Therefore, we integrated biochemical measurements, 16S rDNA sequencing, and transcriptomic analysis to evaluate the impact of 96 h of ammonia nitrogen exposure on oxidative stress, the immune response, lipid metabolism, and gut microbiota in the liver and intestine of lota lota. In this study, lota lota exhibited oxidative damage and immunosuppression following ammonia nitrogen exposure, as evidenced by decreased antioxidant and immune enzyme activities and a concomitant increase in malondialdehyde (MDA) content. Furthermore, ammonia nitrogen exposure altered gut microbial diversity: the abundance of Proteobacteria decreased, whereas that of Bacteroidota and Firmicutes increased. In addition, transcriptomic analysis revealed that the expression of genes related to fatty acid metabolism (fabp, elovl6, pltp) and gut barrier dysfunction (collagen, ecm) was altered. Notably, lipopolysaccharide (LPS) was identified as a key microbial-derived signal triggered by ammonia nitrogen, as demonstrated by ELISA. The translocation of LPS from gut to liver, resulting from compromised intestinal barrier integrity, activated the gut-liver axis and contributed to hepatic lipid metabolic disorders. Overall, the current study elucidates the gut-liver axis-mediated mechanism of ammonia nitrogen toxicity and provides valuable information for understanding the toxic effects of ammonia nitrogen on lota lota and conserving this threatened cold-water species.
The presence of antibiotics in aquatic environments poses risks to aquatic organisms. However, the effects of these antibiotics on freshwater green algae species and their associated metabolic responses are not well understood. This study evaluated the effects of four antibiotics-clarithromycin (CTM), enrofloxacin (ENR), tetracycline (TC), and sulfamethazine (SMZ)-on six algal species: Chlorella pyrenoidosa, Raphidocelis subcapitata, Coelastrum sp., Tetradesmus obliquus, Scenedesmus acuminatus, and Scenedesmus quadricauda. The evaluation utilized Fourier transform infrared spectrometry (FTIR) in combination with metabolomic analysis. The four antibiotics exhibited markedly different toxicity to the six green algae, with an overall toxicity ranking of CTM > TC > ENR > SMZ, corresponding to high (CTM), moderate (TC and ENR), and low (SMZ) toxicity. CTM showed the highest toxicity at the μg/L level and SMZ the lowest at the mg/L level, and EC50 values varied by up to 2-3 orders of magnitude across species. The differing modes of action of the antibiotics likely account for this variance in toxicity among the six algal species. FTIR and metabolomic analyses demonstrated that exposure to the antibiotics altered lipid composition (e.g., CH3/lipid ratio) and protein conformation, and disrupted key metabolic pathways, particularly those related to amino acid metabolism in C. pyrenoidosa. These findings provide highlight the risks posed by antibiotics to freshwater algae and to aquatic ecosystems.
Heavy metal pollution, particularly copper (Cu) and cadmium (Cd), poses a serious threat to aquatic ecosystems due to its toxicity and bioaccumulation potential. This study investigated the effects of individual and combined exposure to Cu and Cd on largemouth bass, focusing on the gut-liver axis and the TLR4/NF-κB pathway in hepatotoxicity. A total of 480 size-matched fish were randomly assigned to four groups (Control, Cu, Cd, and Cu + Cd) and exposed to sublethal concentrations of CuSO₄ (9.275 mg/L) and CdCl₂·2.5H₂O (1.15285 mg/L) for 14 days, followed by physiological, molecular, and omics analyses. Results showed that both single and combined exposures disrupted intestinal structure and barrier function, accompanied by downregulation of tight junction genes. Oxidative stress responses were tissue-specific, with increased reactive oxygen species (ROS) and MDA levels in the intestine but decreased levels in the liver, along with suppressed antioxidant enzyme activities. Inflammatory responses were activated, as indicated by elevated cytokine levels and upregulation of TLR4/MyD88/NF-κB signaling in the liver. Microbiota analysis revealed that Cd exposure increased gut microbial diversity, whereas Cu + Cd co-exposure reduced α-diversity and altered microbial composition, with increased Proteobacteria and decreased beneficial taxa such as Bacteroidota and Firmicutes. Untargeted metabolomics showed that hepatic metabolic profiles were altered, mainly affecting lipid, energy, and immune-related pathways. Overall, combined Cu + Cd exposure exerted stronger toxic effects than single exposures, inducing more severe intestinal damage, microbial dysbiosis, and metabolic disturbances. These findings highlight the critical role of the gut-liver axis in mediating heavy metal toxicity.
Fipronil is a widely detected phenylpyrazole insecticide in aquatic ecosystems that acts as a GABA receptor antagonist in arthropods, but its effects on vertebrate cardiorespiratory physiology are not well understood. In this study, we investigated whether environmentally relevant concentrations of fipronil (0.5 μg l-1, 96 h) impair the cardiovascular and ventilatory responses of rainbow trout (Oncorhynchus mykiss) during aerobic exercise. Using dorsal aortic and buccal cannulations, we continuously recorded heart rate (fH), mean arterial pressure (MAP), ventilatory rate (fV) and amplitude (VAMP) before and after pharmacological blockades (autonomic blockades using atropine and propranolol, and a GABAergic blockade to simulate the effects of fipronil with bicuculline). Exposure to fipronil induced significant tachycardia (∼35%), without affecting MAP or fV. It also increased intrinsic heart rate, even after double autonomic blockade, suggesting direct effects on cardiomyocytes. During exercise, contaminated fish exhibited impaired ventilatory amplitude and autonomic imbalance characterized by vagal withdrawal and sympathetic overdrive. Interestingly, the GABA antagonist bicuculline reproduced the tachycardic pattern observed in contaminated fish, suggesting that fipronil acts through GABAergic disruption of the neural centers involved in cardiac control. These sublethal impairments in autonomic regulation likely reduce the aerobic capacity and ecological resilience of fish in contaminated habitats. ENVIRONMENTAL IMPACT STATEMENT: This study demonstrates that acute exposure to the globally prevalent insecticide fipronil, at an environmentally relevant concentration, causes significant sublethal dysfunction in the neurophysiological control of cardiorespiratory systems in rainbow trout. By disrupting GABAergic signaling, fipronil compromises the integrated cardiovascular and ventilatory responses essential for sustaining aerobic exercise. These findings reveal a critical pathway through which pesticide pollution can impair physiological performance, thereby reducing the fitness and adaptive resilience of fish populations facing concurrent anthropogenic stressors such as climate change and habitat degradation. The results underscore the need to consider subtle, neurotoxic mechanisms in environmental risk assessments for aquatic ecosystems.
This study investigates the photoexcited larvicidal toxicity of zinc oxide (ZnO) nanoparticles against Aedes aegypti larvae under ultraviolet A (UV-A) irradiation. Three ZnO samples with different particle sizes were prepared from a colloidal suspension, with the smallest fraction (ZnO-C1) exhibiting an average hydrodynamic diameter of 70.85 nm, while the largest reached 3250 nm. No larval mortality was observed under dark conditions. In contrast, acute toxicity occurred only upon UV-A exposure, revealing a significant light-dependent toxic response that has been less explored in the context of specific ZnO size fractions. Larvicidal bioassays conducted according to World Health Organization guidelines demonstrated a remarkably low LC₅₀ value of 2.50 mg/L for ZnO-C1, among the lowest reported for ZnO-based larvicidal activity. FESEM-EDX analysis detected zinc within the larval body and midgut, indicating nanoparticle uptake through ingestion and respiration. Mechanistic evaluation suggests that UV photoexcitation generates electron-hole pairs in ZnO, promoting the formation of reactive oxygen species (ROS) and Zn2+ ions that cause membrane disruption and internal tissue damage. The enhanced toxicity of ZnO-C1 is attributed to its smaller size, larger surface area, and improved dispersion stability, which facilitate greater ROS generation and ionic dissolution. These findings demonstrate a distinct UV-dependent mode of action and highlight the potential of nanoscale ZnO as a photoactivated larvicidal agent against Ae. aegypti.
