Polyfluoroalkyl phosphate esters (PAPs) are a class of emerging per- and polyfluoroalkyl substances, while their bacterial toxicity is lacking. Herein, the wild-type (WT) and Δacriflavine resistance B (acrB) strains of E. coli K12 were exposed to 6:2 diPAP, one frequently detected PAP, respectively. The intracellular concentrations of 6:2 diPAP increased in the absence of acrB. At the individual level, 1 nmol L-1 6:2 diPAP significantly inhibited the growth and elevated reactive oxygen species levels of the WT strain. In the absence of acrB, such inhibitory and stimulating effects were more pronounced at 0.1 and 1 nmol L-1 6:2 diPAP compared with the WT strain. At the population level, biofilm formation was promoted in the WT strain while was inhibited in the ΔacrB strain at 0.1 and 1 nmol L-1 6:2 diPAP. The expressions of S-ribosylhomocysteine lyase, motility protein A and ybaJ followed the same trend as biofilm formation, while those of tryptophanase and haemolysin expression-modulating protein showed opposite trends. The results demonstrate that 6:2 diPAP affected biofilm formation by regulating their expressions. In accordance with biofilm formation, glutathione (GSH) metabolism was activated in the WT strain while was suppressed in the ΔacrB strain by 1 nmol L-1 6:2 diPAP. The defect in biofilm formation of the ΔacrB strain was partly rescued by the addition of four metabolites related with GSH metabolism, consolidating the involvement of GSH metabolism in biofilm formation. Our study shows the indispensable role of acrB in the toxicities of 6:2 diPAP to E. coli at environmentally relevant concentrations.
Isothiazolinone biocides are increasingly recognized as emerging contaminants in aquatic environments. Among them, 2-n-octyl-4-isothiazolin-3-one (OIT) and 4,5-dichloro-2-n-octyl-4-isothiazolin-3-one (DCOIT) may co-occur in receiving waters, yet their combined effects on marine organisms remain poorly understood. The Mediterranean mussel Mytilus galloprovincialis was exposed for 14 days to OIT (2.81 µg L⁻¹), DCOIT (1 µg L⁻¹), and their mixture (MIX), selected to represent environmentally relevant exposure levels. Cellular viability, regulatory volume decrease, haemocyte phagocytic activity, and the expression of oxidative stress-, xenobiotic metabolism-, and inflammation-related genes were evaluated. Overall cell viability remained high, although OIT significantly reduced haemocyte membrane integrity and lysosomal stability. All exposure conditions altered cell volume regulation and markedly reduced haemocyte phagocytic activity, with the strongest reduction observed under co-exposure. Transcriptional analyses revealed tissue-wide modulation of MnSOD, Cu/ZnSOD, CYP4Y1, TNF-α, and IL-17, with the MIX group generally showing the greatest responses. In the digestive gland, CYP4Y1 expression reached a 4.13-fold increase, while MnSOD and TNF-α increased 3.33- and 3.50-fold, respectively, compared with controls. Co-exposure to OIT and DCOIT at low concentrations was associated with integrated cellular, osmoregulatory, immune, and transcriptional responses in M. galloprovincialis. These findings indicate that limited cytotoxicity does not exclude substantial sublethal effects and highlight the ecological relevance of assessing isothiazolinones as co-occurring contaminants rather than solely as individual compounds.
To counter the inefficiency of conventional pesticides, nanopesticides have emerged as a promising alternative to enhance delivery and increase target specificity. However, the unique set of physicochemical properties that promise to enhance their efficacy also raises concerns about their environmental behaviour and toxicity. Among these, copper oxide nanoparticles (CuONPs) are widely used as an antimicrobial agent. While studies focus on their properties against pathogens, their effects on non-target fungal communities and broader ecological functions remain largely unknown. Our study aims to evaluate the impacts of CuONPs on freshwater fungal communities and the key ecosystem functions they support. Specifically, we assessed (1) host-parasite interactions and disease transmission using a Daphnia-microparasitic yeast experimental system (disease model), and (2) organic matter decomposition mediated by aquatic hyphomycetes (decomposition model). Results reveal contrasting and process-specific effects. CuONPs decreased disease prevalence (IC50 = 0.30 mg l-¹) but impaired host fitness through reduced survival and fecundity. In contrast, organic matter decomposition was functionally resistant, with stable litter mass loss and fungal biomass masking other effects, including the inhibition of fungal reproduction (IC50 = 2.53 mg l-¹) and a marked restructuring of the decomposer community (based on conidial morphology). Collectively, our findings demonstrate that CuO nanoparticles can subtly reshape ecological interactions and community composition. From a management perspective, this highlights the need of incorporating species interactions, community-level endpoints, and functional responses (disease dynamics, decomposition) into environmental risk assessments of contaminants of emerging concern.
Zebrafish exhibit innate phototaxis, making light stimulation a promising tool for water toxicity monitoring.This study systematically investigated the effects of light color (red, yellow, green, blue), intensity (0-2400 lx), and stimulation duration (1-3 min) on zebrafish phototactic and locomotor behaviors, and established an optimized photobehavioral assay. The results showed that zebrafish exhibited strong positive phototaxis toward red light (612 nm, peak 78.74%) and clear negative phototaxis toward blue light (450 nm, peak -36.23%), with peak behavioral responses at 700 lx and 300 lx, respectively. An optimized intermittent stimulation protocol (red light: 2 min stimulation / 15 min interval; blue light: 1 min stimulation / 15 min interval) effectively maintained behavioral sensitivity while minimizing habituation. A “startle-phototaxis-adaptation” behavioral sequence was identified, consisting of an initial luminance-driven startle (first minute) followed by wavelength-specific phototaxis. Validation with Cd2+ exposure revealed concentration-dependent suppression of phototaxis: at 2 TU, the red-light phototaxis rate decreased from 78.9% to -55.9%. With high repeatability, this photobehavioral assay acts as a sensitive, ecologically meaningful proof-of-concept tool, using Cd2+ as a model toxicant to support subsequent aquatic pollutant risk evaluation.
Per- and polyfluoroalkyl substances (PFAS) are persistent and bioaccumulative contaminants of emerging concern in aquatic ecosystems, yet their uptake dynamics and physiological effects on macrophytes remain poorly understood. This study investigated PFAS bioaccumulation and physiological responses of Lemna minor exposed to seven PFAS compounds under both acute and chronic conditions. Longer exposure durations resulted in detectable accumulation of a greater number of PFAS, particularly at lower exposure concentrations. Concentration-dependent accumulation was observed, although the direction and magnitude of these relationships varied among PFAS types. PFBS showed the highest accumulation, with concentrations up to 8736 ± 5715 ng/g dw detected after 10 days of exposure to 100,000 ng/L. When expressed as bioconcentration factors (BCFs), an inverse relationship with exposure concentration indicated higher uptake efficiency at environmentally relevant concentrations. The highest BCF (6076 ± 898 L/g) was observed for PFBS after 20 days of exposure to 1 ng/L. Differences in accumulation and BCF values were attributed to functional group and chain length, which affect solubility and hydrophobicity. Despite measurable bioaccumulation, physiological responses were generally minor, as neither growth nor photosynthetic efficiency (Fv/Fm and Y(II)) showed consistent impairment. Growth stimulation was observed in a limited number of treatments, suggesting potential hormetic effects. Overall, these findings highlight that both exposure duration and concentration range should be considered when assessing PFAS bioaccumulation in macrophytes, and that integrating time-dependent accumulation into risk assessments is essential for accurately evaluating PFAS behaviour and ecological risks.
Trimethoprim (TMP) and sulfamethoxazole (SMX) are commonly detected antibiotics in marine environments, yet their potential hazards to marine microalgae under co-occurrence remain poorly understood. This study investigated the combined toxic effects of environmentally relevant concentrations of TMP (200 ng/L) and SMX (50-500 ng/L) on two marine microalgae species, Skeletonema costatum and Phaeodactylum tricornutum. The results revealed that the antibiotic mixture stimulated the growth of P. tricornutum. In the group treated with 100 ng/L SMX plus 200 ng/L TMP, algal cell density increased significantly compared with the control group (P < 0.05), with a growth rate 14.2% higher than the control. By contrast, the combined antibiotics had no notable impact on the growth of S. costatum. The mixture also altered the chlorophyll content, cell membrane hydrophobicity and antioxidant defense system of P. tricornutum. After 144 h of exposure to 250 ng/L SMX and 200 ng/L TMP, the malondialdehyde (MDA) content in P. tricornutum rose significantly (P < 0.05), with MDA content reaching 175.46% of the control level. At the molecular level, the activation of the ABC transporters pathway served as an adaptive detoxification strategy for microalgae under single TMP stress. The combined transcriptional and metabolic changes in the valine, leucine and isoleucine biosynthesis pathway were the key factor for TMP to amplify SMX toxicity. The combined disruption of this pathway, porphyrin metabolism and lipid metabolism jointly affected algal growth. The findings provide a reference for evaluating the potential risks of mixed antibiotics to marine primary productivity.
Organophosphate ester flame retardants and nanoplastics (NPs) frequently co-occur in aquatic environments; however, their combined effects on fish visual function remain poorly understood. Here, we investigated whether NPs modulate triphenyl phosphate (TPhP)-induced ocular toxicity and color-guided behavior in zebrafish. Fish were exposed to environmentally relevant concentrations of TPhP, NPs, or their mixture (TNP), followed by color preference assays, retinal histopathology, apoptosis analysis, and eye-tissue transcriptomics with targeted gene validation. TPhP exposure significantly disrupted green and red color preferences and altered color-dependent spatial distribution. These behavioral deficits were accompanied by retinal structural damage, including thinning of the inner neuronal and photoreceptor layers, downregulation of opsin-related genes, and increased apoptosis. In contrast, these alterations were partially attenuated under co-exposure conditions. Transcriptomic analyses further revealed distinct molecular signatures under single exposures; TPhP predominantly affected pathways associated with retinal structure, energy metabolism, and junctional integrity, whereas NPs primarily activated inflammation- and cell death-related processes. Co-exposure elicited a broader and more complex transcriptional response, characterized by coordinated reprogramming of immune-inflammatory, metabolic, tight junction, and regulated cell death pathways, suggesting a non-additive interaction between TPhP and NPs. Collectively, these findings demonstrate that NPs reshape TPhP-induced ocular toxicity through complex transcriptional reprogramming rather than simply enhancing or alleviating individual toxic effects. This study highlights the importance of considering mixture-induced molecular adaptation when evaluating the ecological risks of emerging contaminants and supports color-guided behavior as a sensitive endpoint for assessing visual toxicity in aquatic organisms.
Freshwater ecosystems are exposed to a wide range of pollutants, including veterinary pharmaceuticals. Among those pharmaceuticals, ivermectin, an antiparasitic compound, has received increasing attention over the last decades due to its high ecotoxicity, with effects being reported in the sub ng/L range. While several orders have been investigated for their sensitivity to ivermectin, Trichoptera, a species-rich and ecologically significant insect order, have largely been overlooked. Here we assessed acute (7 days) and sub-chronic (5 weeks) (mortality, feeding rate, growth and fatty acid (FA) profile) effects induced by ivermectin in Chaetopteryx villosa, a common Trichoptera species in central Europe. Tested ivermectin concentrations ranged from 10 to 10,000 ng/L and 75 to 2,400 ng/L in case of the acute and sub-chronic toxicity test, respectively. LC₅₀ values declined from 2.81 µg/L after 7 days of exposure to 0.74 µg/L after five weeks. Moreover, ivermectin exposure resulted in a strong concentration-dependent reduction in growth assessed by changes in their dry weight (i.e., of 60 % at 300 ng/L, 54 % at 600 ng/L, 100 % at 1,200 ng/L compared to the ivermectin-free control), an observation likely driven by a significantly reduced feeding activity. Moreover, concentrations of individual FAs in C. villosa larvae were significantly reduced at 300 and 600 ng/L, while lower concentrations tended to increase concentrations of several FAs. Similarly, ivermectin significantly affected FA profile and the relative contribution of saturated, monounsaturated and polyunsaturated FAs. Overall, our results demonstrate concentration- and time-dependent effects of ivermectin on key life-history traits of C. villosa potentially leading to effects in aquatic food webs through the shift in FA profiles.
Research on the ecotoxicity of biodegradable microplastics (MPs) in Daphnia magna is rapidly growing, while their population-level effects remain unclear. This study compared the population-level effects of conventional MPs (polyethylene terephthalate, PET) and biodegradable MPs (polyhydroxybutyrate, PHB) on D. magna over 45 days. Gut microbiome and transcriptomic analyses were also conducted to elucidate the underlying mechanisms of the observed responses. PET MPs significantly (p < 0.05) decreased D. magna population biomass relative to control, whereas PHB MPs showed no significant effects. Both types of MPs altered the gut microbial community structure of D. magna, with PHB inducing a selective increase in potential degraders. Transcriptomic data showed that PET MPs significantly (p < 0.05) increased the expression of genes related to stress and defense responses. In contrast, PHB MPs significantly (p < 0.05) upregulated genes associated with metabolic processes. Further research incorporating direct assessment of PHB degradation, host energy assimilation, and wider range of exposure concentrations is required to clarify the mechanisms underlying the distinct responses to biodegradable and conventional MPs. This study highlights the importance of considering polymer types in MP environmental risk assessments and underscores the need of integrating multiple endpoints for a comprehensive evaluation.
Nonsteroidal anti-inflammatory drugs are among the most commonly used pain-relieving, anti-inflammatory and antipyretic drugs. When they were introduced to the pharmaceutical market, large-scale toxicological studies were conducted to determine their effect on the human body. However, their widespread use contributed to the presence of these drugs and their metabolites in the environment, including soil and water, where they can affect organisms that are not the direct targets of action. Hence, it is extremely important to correctly estimate the toxicity of these drugs on organisms inhabiting various biocenoses. The presented review attempts to summarise the current results on the effect of nonsteroidal anti-inflammatory drugs obtained in studies on model organisms and presents a wider spectrum of issues related to the toxicity of these drugs, including the human microbiome. The paper shows that limiting studies only to typical model organisms, despite the fact that such studies provide a number of answers, may underestimate the negative impact of these compounds in relation to other, less frequently tested microorganisms.
Given the increasing co-occurrence of pharmaceuticals and personal care products (PPCPs) and eutrophication in aquatic environments, it is crucial to understand PPCP-nutrient-plant interactions and their combined ecological impact. However, the literature regarding quantitative thresholds and metabolic trade-offs remain highly fragmented. In this meta-analysis, we compiled a dataset comprising 407 observations from 38 rigorously screened studies, covering various PPCP classes and floating, submerged, and emergent plant habitats. Remediation efficiency and physiological stress markers specifically enzymatic defense and oxidative damage were selected as informative endpoints for assessing plant responses to combined exposure. The combined stressors' phytotoxicity generally manifested as severe oxidative stress and metabolic exhaustion in submerged species, despite high apparent initial removal. However, the observed remediation phenotype was heavily influenced by experimental conditions, particularly exposure concentration, transitioning from a metabolic primer to an inhibitory stressor above a 10 mg/L threshold. Conversely, physiological stability and active removal efficiency were maintained in floating species and specific elite emergent candidates, particularly Vetiveria zizanioides, following exposure to co-contamination. Ultimately, this meta-analysis highlights the severe metabolic costs aquatic plants face in co-contaminated systems. We identified a critical toxicity tipping point at a 10 mg/L exposure concentration. Beyond this threshold, we observed a critical performance shift at a 10 mg/L exposure concentration. We hypothesize that beyond this threshold, plant defense systems are severely overwhelmed, and the observed removal of contaminants may transition from active biological degradation to passive physical sorption. These findings provide essential mechanistic insights into how combined chemical and nutrient stressors disrupt plant physiology, which is critical for understanding their broader ecological impact.
Microplastic (MP) pollution poses increasing ecological risks to marine filter feeders, yet how different MP characteristics collectively influence biological toxicity and environmental redistribution remains poorly understood. In this study, the mussel Mytilus coruscus was exposed to two representative MPs, laboratory-model polyethylene (PE) microbeads and environmentally relevant and polyethylene terephthalate (PET) microfibers, to investigate particle fate, tissue distribution, energy metabolism, and antioxidant responses. Mussels rapidly removed >90% of suspended MPs from the water column within 72 h, facilitating particle transfer to tissues and sediments. However, the two MPs induced markedly different distribution patterns and physiological responses. PET microfibers showed substantially greater retention in gill tissues (∼30%) than PE microbeads (<10%), accompanied by reduced clearance efficiency and persistent oxidative stress characterized by elevated lipid peroxidation after depuration. In contrast, PE microbeads mainly induced metabolic compensation through enhanced ETS activity and mobilization of energy reserves. Tissue-specific responses further revealed functional differentiation between gills and digestive glands in particle processing and physiological regulation. The results demonstrate that combined differences in MP morphology, composition, and structure are likely associated with distinct environmental fates and organismal stress strategies. These findings provide new insight into how environmentally realistic MPs shape tissue-specific physiological strategies in marine filter feeders and highlight the importance of particle heterogeneity in MP ecotoxicology.
Microplastic pollution in marine environments has raised significant concerns. However, it remains unclear whether bioplastic particles are indeed more environmentally-friendly and less noxious to marine species than conventional petroleum-based plastics. This study aimed to adopt a holistic approach (integrating animal behaviour, plasma metabolite profiling, brain histopathology and biochemical markers indicative of neural function and antioxidant responses) to assess the ecotoxicological effects of juvenile gilthead seabream (Sparus aurata) orally exposed to petroleum-based (polyethylene terephthalate, PET), and bio- (polybutylene succinate, PBS and polypropylene 2,5-furandicarboxylate, PPF) microplastics, for a period of 28 days. The ability of fish to recover from stress induced by plastics exposure was also investigated, by analysing the aforementioned endpoints following a seven-day depuration period. Of the twelve plasmatic metabolites assessed, only two were significantly affected by microplastics ingestion - glucose levels increased in fish exposed to PET, while amylase activity decreased after exposure to all three microplastics. There was no evidence of severe and/or irreversible alterations in neuronal function (i.e. acetylcholinesterase activity), cortisol levels, behaviour or brain histopathological structure. The antioxidant defence system was mildly affected by the exposure to PET and PBS (i.e. decreased catalase activity), although this effect was no longer observed during the depuration period. At the end of the exposure period no cell lipid peroxidation occurred, however, PPF stimulated the ubiquitin-proteasome pathway (i.e. increased ubiquitin content). Overall, the exposure to these microplastics could not be conclusively or clearly linked to persistent toxicity in gilthead seabream.
Plastic pollution is one of the major global concerns. The replacement of traditional synthetic polymers by “greener” alternatives (i.e., biodegradable and/or bio-based plastics) with lower toxicity and environmental impact has been identified as a key strategy to reduce the environmental hazards posed by plastics. Despite the improvements brought by bioplastics from the environmental degradability and overall life-cycle standpoint, a few recent studies have reported that these new polymers have similar (or, even, higher) toxicity to conventional plastics (such as PET), primarily due to their high biodegradability and the subsequent leaching of additives that compose them. Hence, the present review aims to summarize the ecotoxicological effects of some currently used bioplastics and green plasticizers on aquatic organisms, and to compare them with those reported for conventional fossil-fuel-based plastics. The adverse effects observed upon exposure to bioplastics and their additives include impaired survival rates, as well as development and histopathological alterations in various organs (liver, gills, digestive system, and gonads), biochemical and molecular evidence of oxidative stress, altered immunity, and impaired reproduction. The scarcity of empirical toxicological studies and the current lack of human exposure recommendations urgently call for further research efforts on commercially available bioplastics (and their additives). Such efforts are needed to determine whether these materials represent a real eco-friendly alternative, as well as to develop new ones that meet both the industrial, environmental and toxicological requirements. This review aims to help identifying the most promising biopolymer alternatives from an ecotoxicological perspective, while highlighting on the current research gaps.
The global phase-out of legacy brominated flame retardants (BFRs) has coincided with the increasing detection of novel BFRs in the environment, creating a complex scenario of coexisting legacy and novel pollutants. Aquatic systems serve as major sinks for BFRs, making it imperative to assess the potential ecological risks posed by BFRs to aquatic animals. This study synthesized 8009 valid data points from 137 publications using meta-analysis, machine learning, and the Geodetector model. Meta-analysis revealed that biological factors (life stage and ecological niche) are important determinants of species sensitivity to BFRs, with early life stages, adults and benthic animals exhibiting stronger responses. Exposure conditions (BFR type, exposure time and concentration) were also important factors driving toxicity, particularly the significant effects induced by banned BFRs, high concentration exposure (> 1000 μg/L) and chronic exposure (> 14 days). A non-monotonic dose-response relationship was observed, indicating that the toxic effects did not follow a simple linear concentration‑dependent pattern. Toxicological endpoint analysis identified cytotoxicity and endocrine disruption as dominant effects. Machine learning and Geodetector model identified endpoint and BFR type as the most important factors, with widespread interactive enhancement effects among the factors. Notably, decabromodiphenyl ethane (DBDPE), a primary alternative to decabromodiphenyl ether (BDE-209), exhibited effect magnitudes comparable to or even greater than those of BDE‑209 for certain endpoints (e.g., programmed cell death and thyroid endocrine). Its toxicity was also more susceptible to modulation by exposure conditions. These findings challenge the perceived safety of DBDPE as a "safer alternative" and underscore its non‑negligible ecological risk. Mechanistically, comparative analysis of ecological niches across species indicated that the same BFR could trigger divergent adverse outcome pathways in different species, attributable to species-specific molecular initiating events.
Bisphenol F (BPF) has been widely used as a major substitute for bisphenol A (BPA) in numerous consumer and industrial products. Its environmental presence is increasingly documented, with frequent detections in surface water, sediment, and sewage sludge across various countries, often at notably high detection rates and concentrations. BPF exhibits a range of adverse effects, including developmental toxicity, neurotoxicity, oxidative stress, and endocrine‑disrupting activity. It also disrupts the reproductive and endocrine systems by altering the metabolism or synthesis of endogenous hormones or through more complex epigenetic mechanisms. Given that BPF induces multiple toxicities, including effects on developing germ cells, such epigenetic alterations in the germline genome may transmit harmful consequences to subsequent generations. In this review, we summarize the reported concentrations and detection of BPF in the aquatic environments, followed by a review of the literature on its multifaceted toxicity of BPF exposure. We aim to provide a comprehensive assessment of its potential ecological and organismal health risks. Nevertheless, significant knowledge gaps remain. Future studies should prioritize environmentally relevant chronic exposure, mixture toxicity, identification of BPF‑specific biomarkers, and multigenerational ecological impacts.
Decomposition of organic matter is critical to the biogeochemical cycling of carbon and nutrients in all ecosystems. In streams, decomposition can be influenced by anthropogenic impacts, including contaminants. Per- and polyfluoroalkyl substances (PFAS) are resistant to degradation and widespread in freshwater ecosystems, yet little is known about their influence on organic matter processing in streams. We paired an observational field study of a PFAS-impacted stream via food processing wastewater with a 28-day laboratory experiment to investigate PFAS effects on leaf litter processing using multiple lines of evidence. Leaf litter decomposition rates and microbial respiration were significantly lower downstream of the PFAS point-source compared to the paired upstream site. We hypothesized that PFAS suppressed microbial activity thereby resulting in slower decomposition rates. To investigate further and better isolate PFAS, we dosed aquatic mesocosms containing leaf packs with differing concentrations of perfluorooctane sulfonate (PFOS). Over the 28-day incubation, decomposition rates did not differ, but we observed lower respiration rates on day 28 and higher dissolved organic carbon (DOC) in PFOS-dosed mesocosms, suggesting that PFAS may suppress microbial activity and inhibit carbon processing. These results underscore that concerns regarding PFAS contamination extend beyond organismal toxicity to ecosystem-level effects. Further research is needed to understand the scale, implications, and mechanisms responsible for these changes.
