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The broadly utilized biocide triclosan (TCS) is continuously discharged in water compartments worldwide, where it is detected at concentrations of ng-µg/L. Given its lipophilicity and bioaccumulation, TCS is considered potentially harmful to human and environmental health and also as a potential endocrine disruptor (ED) in different species. In aquatic organisms, TCS can induce a variety of effects: however, little information is available on its possible impact on invertebrate development. Early larval stages of the marine bivalve Mytilus galloprovincialis have been shown to be sensitive to environmental concentrations of a number of emerging contaminants, including EDs. In this work, the effects of TCS were first evaluated in the 48 h larval assay in a wide concentration range (0.001-1,000 μg/L). TCS significantly affected normal development of D-veligers (LOEC = 0.1 μg/L; EC 50  = 236.1 μg/L). At selected concentrations, the mechanism of action of TCS was investigated. TCS modulated transcription of different genes involved in shell mineralization, endocrine signaling, ceramide metabolism, and biotransformation, depending on larval stage (24 and 48 h post-fertilization-hpf) and concentration (1 and 10 μg/L). At 48 hpf and 10 μg/L TCS, calcein staining revealed alterations in CaCO 3 deposition, and polarized light microscopy showed the absence of shell birefringence due to the mineralized phase. Observations by scanning electron microscopy highlighted a variety of defects in shell formation from concentrations as low as 0.1 μg/L. The results indicate that TCS, at environmental exposure levels, can act as a developmental disruptor in early mussel larvae mainly by interfering with the processes of biomineralization., (© 2023. The Author(s).)

Marine biofouling pollution is a process that impacts ecosystems and the global economy. On the other hand, traditional antifouling (AF) marine coatings release persistent and toxic biocides that accumulate in sediments and aquatic organisms. To understand the putative impact on marine ecosystems of recently described and patented AF xanthones (xanthones 1 and 2), able to inhibit mussel settlement without acting as biocides, several in silico environmental fate predictions (bioaccumulation, biodegradation, and soil absorption) were calculated in this work. Subsequently, a degradation assay using treated seawater at different temperatures and light exposures was conducted for a period of 2 months to calculate their half-life (DT 50 ). Xanthone 2 was found to be non-persistent (DT 50  < 60 days) at 50 μM, contrary to xanthone 1 (DT 50  > 60 days). To evaluate the efficacy of both xanthones as AF agents, they were blended into four polymeric-based coating systems: polyurethane- and polydimethylsiloxane (PDMS)-based marine paints, as well as room-temperature-vulcanizing PDMS- and acrylic-based coatings. Despite their low water solubility, xanthones 1 and 2 demonstrated suitable leaching behaviors after 45 days. Overall, the generated xanthone-based coatings were able to decrease the attachment of the Mytilus galloprovincialis larvae after 40 h. This proof-of-concept and environmental impact evaluation will contribute to the search for truly environmental-friendly AF alternatives., (© 2023. The Author(s).)

Organotin-based (OTs: TBT and TPT) antifouling paints have been banned worldwide, but recent inputs have been detected in tropical coastal areas. However, there is a lack of studies evaluating the toxicity of both legacy and their substitute antifouling booster biocides (e.g., Irgarol and diuron) on neotropical species. Therefore, the acute toxicity of four antifouling biocides (TBT, TPT, Irgarol, and diuron) was investigated using the marine planktonic organisms Acartia tonsa and Mysidopsis juniae, the estuarine tanaid Monokalliapseudes schubarti (water exposure), and the burrowing amphipod Tiburonella viscana (spiked sediment exposure). Results confirmed the high toxicity of the OTs, especially to planktonic species, being about two orders of magnitude higher than Irgarol and diuron. Toxic effects of antifouling compounds were observed at levels currently found in tropical coastal zones, representing a threat to planktonic and benthic invertebrates. Furthermore, deterministic PNEC marine sediment values suggest that environmental hazards in tropical regions may be higher due to the higher sensitivity of tropical organisms. Since regulations on antifouling biocides are still restricted to a few countries, more ecotoxicological studies are needed to derivate environmental quality standards based on realistic scenarios. The present study brings essential contributions regarding the ecological risks of these substances in tropical and subtropical zones., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Since the banning of tributyltin, the addition of inorganic (metal oxides) and organic (pesticides, herbicides) biocides in antifouling paint has represented an unavoidable step to counteract biofouling and the resulting biodeterioration of submerged surfaces. Therefore, the development of new methods that balance antifouling efficacy with environmental impact has become a topic of great importance. Among several proposed strategies, natural extracts may represent one of the most suitable alternatives to the widely used toxic biocides. Posidonia oceanica is one of the most representative organisms of the Mediterranean Sea and contains hundreds of bioactive compounds. In this study, we prepared, characterized, and assessed a hydroalcoholic extract of P. oceanica and then compared it to three model species. Together, these four species belong to relevant groups of biofoulers: bacteria (Aliivibrio fischeri), diatoms (Phaeodactylum tricornutum), and serpulid polychaetes (Ficopomatus enigmaticus). We also added the same P. oceanica extract to a PDMS-based coating formula. We tested this coating agent with Navicula salinicola and Ficopomatus enigmaticus to evaluate both its biocidal performance and its antifouling properties. Our results indicate that our P. oceanica extract provides suitable levels of protection against all the tested organisms and significantly reduces adhesion of N. salinicola cells and facilitates their release in low-intensity waterflows., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Eco-toxicity profiles for commonly used disinfectants were lacking. Available traditional toxicity techniques have some limitations (assessments and ethical issues). Behaviour toxicology is a promising research area towards early warning and non-invasive approaches. We studied the potential eco-toxic effects of sodium hypochlorite (NaOCl) on the swimming behaviour of zebrafish. Zebrafish were exposed to different concentrations (Treatment I, Treatment II, Treatment III, and Treatment IV) of NaOCl for 360 h. Recovery study (144 h) was conducted for NaOCl treatment groups. The swimming behaviour of zebrafish was quantified efficiently using an online monitoring system (OMS). OMS dataset was processed for determination of behavioural differences by MATLAB and SPSS. Compared to the control group, the swimming strength of zebrafish under NaOCl treatments declined significantly (p < 0.001). Avoidance behaviour has occurred on zebrafish under NaOCl exposure periods. Furthermore, NaOCl toxicity also adjusted circadian rhythms on zebrafish. Zebrafish swimming strength was significantly (p < 0.001) improved under-recovery periods. Moreover, normal diurnal patterns have occurred. NaOCl could cause behavioural abnormalities in non-target organisms. Continuous exposure to common disinfectants could cause external and internal stress on non-target organisms, resulting in behavioural changes and circadian rhythm adjustments. Continuous changes in behavioural and circadian rhythms might reduce organisms' fitness and adaptation capacity. This study highlights (1) the importance of computer-based toxicity assessments, and (2) swimming behaviour is an early warning biomarker for eco-toxicity studies., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Algal organic matter (AOM) in water reservoirs is a worldwide concern for drinking water treatment; once it is one of the main precursors for disinfection by-products formation (DBPs). In this context, this study investigated the ecotoxicity of DBPs from chlorination of AOM to Ceriodaphnia silvestrii and Daphnia similis (Crustacea, Cladocera). The bioassays evaluated three scenarios, including the AOM extracted from Chlorella sorokiniana, the quenching condition used in the tests, and the DBPs formed after the chlorination of the two test waters with AOM (with and without bromide presence). The results showed that AOM has no toxic effects for the tested species under typical environmental concentration (5 mg∙L -1 ). However, since AOM is a potential precursor of DBPs, the toxicity of two test waters (TW-1 and TW-2) after the chlorination process (25 mg Cl 2 ·L -1 , for 7 days, at 20 °C) was tested. The sample with higher toxicity to the tested species was TW-1, in which chloroform and chloral hydrate were quantified (615 and 267 µg∙L -1 , respectively). However, TW-2 showed lower concentration of chloroform and chloral hydrate (260 and 157 µg∙L -1 , respectively), although bromodichloromethane, dibromochloromethane, and bromoform were also detected (464, 366, and 141 µg∙L -1 , respectively). Although free chlorine is highly toxic to the tested species, the quenching conditions also affected the organisms' survival due to the use of ascorbic acid and the presence of reaction intermediates. Nonetheless, both species were more affected by TW-1 and TW-2 than the quenching condition. These results endorse the importance of removing the AOM before the disinfection process to avoid the formation of DBPs. In addition, ecotoxicological analyses could provide a more comprehensive assessment of water quality, especially considering the challenges of quantifying DBPs and other emerging contaminants., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Recent advances in nanotechnology have allowed the encapsulation of hazardous antifouling (AF) biocides in silica mesoporous nanocapsules (SiNC) reducing their short-term toxicity. However, the chronic effects of such novel nanoadditives remain understudied. The present study aimed to assess short- and long-term sub-lethal effects of soluble forms (DCOIT and Ag) and nanostructured forms (SiNC-DCOIT and SiNC-DCOIT-Ag) of two AF biocides and the "empty" nanocapsule (SiNC) on juveniles of Crassostrea gigas after 96 h and 14 days of exposure. Juvenile oysters exposed for a short period to free DCOIT and AgNO 3 presented worse physiological status comparing with those exposed to the nanostructured forms. The long-term exposure to DCOIT and Ag + caused an extensive biochemical impairment comparing with the tested nanomaterials, which included oxidative damage, activation of the antioxidant defense system, and neurotransmission impairment. Despite the negative effects mostly observed on the health condition index and AChE, the encapsulation of the abovementioned AF biocides into SiNC seems to be a technological advantage towards the development of AF nanoadditives with lower long-term toxicity comparing with the soluble forms of such biocides., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Antifouling paints incorporate biocides in their composition seeking to avoid or minimize the settlement and growing of undesirable fouling organisms. Therefore, biocides are released into the aquatic environments also affecting several nontarget organisms and, thus, compromising ecosystems. Despite global efforts to investigate the environmental occurrence and toxicity of biocides currently used in antifouling paints, the specific active ingredients that have been used in commercial products are poorly known. Thus, the present study assessed the frequencies of occurrence and relative concentrations of biocides in antifouling paint formulations registered for marketing worldwide. The main data were obtained from databases of governmental agencies, business associations, and safety data sheets from paint manufacturers around the world. The results pointed out for 25 active ingredients currently used as biocides, where up to six biocides have been simultaneously used in the examined formulations. Cuprous oxide, copper pyrithione, zinc pyrithione, zineb, DCOIT, and cuprous thiocyanate were the most frequent ones, with mean relative concentrations of 35.9 ± 12.8%, 2.9 ± 1.6%, 4.0 ± 5.3%, 5.4 ± 2.0%, 1.9 ± 1.9%, and 18.1 ± 8.0% (w/w) of respective biocide present in the antifouling paint formulations. Surprisingly, antifouling paints containing TBT as an active ingredient are still being registered for commercialization nowadays. These results can be applied as a proxy of biocides that are possibly being used by antifouling systems and, consequently, released into the aquatic environment, which can help to prioritize the active ingredients that should be addressed in future studies., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Biofouling is a costly burden for the shipping industry. Metal-based antifouling paints are widely used to protect submerged surfaces, but the release of metals from coatings and the recoating of hulls can leach large amounts of copper and zinc into aquatic environments, posing a risk for aquatic ecosystems and biodiversity. With this study, we studied the time-course metal accumulation and oxidative stress in the digestive gland and the gills of Hyriopsis bialatus, an Asian freshwater mussel, exposed to sublethal concentrations of cuprous chloride (50 and 5 µg/L) and zinc sulfate (1000 and 100 µg/L). Time-dependent accumulation was observed after exposure to copper, but zinc uptake was negligible. Integrated biomarker response (IBRv2) and statistical analysis of individual biomarker levels showed a greater biomarker response in the digestive gland and the gills after exposure to the higher concentration of CuCl and ZnSO 4 . Both compounds elicited a biochemical response, especially in the digestive gland. Glutathione peroxidase activity was increased after exposure to both metals at both concentrations, suggesting a powerful defense against lipid peroxidation. The biological impact of zinc was less than that of copper, suggesting mitigated ecological pressure., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

The Pacific oyster (Crassostrea gigas) is an important species in oyster farming worldwide, including in Japan. Hiroshima Bay is one of the most important oyster farming areas in Japan. We investigated the occurrence of antifouling biocides used worldwide including diuron, Irgarol 1051 (Irgarol), and 4,5-dichloro-2-n-octyl-4-isothiazolin-3-one (DCOIT), which have been detected at sub-ppb levels in seawater in Japan, and estimated their no observed effect concentrations (NOECs). In recent years, the spat settlement of Pacific oysters has become poor, which presents a challenge for oyster aquaculture in Hiroshima Bay; hence, we conducted embryotoxicity and larva settlement tests using Pacific oysters. Compared to diuron and Irgarol, DCOIT exhibited a higher toxicity toward oyster embryos, and the minimum 24-h NOEC toxicity value for the oyster embryos was <3 ng/L. The highest concentrations of diuron, Irgarol, and DCOIT in the environmental seawater in the Seto Inland Sea were 27.6, 3.2, and 24 ng/L, respectively. Considering the NOECs, the environmental concentrations of these biocides suggest that the ecological risks posed by diuron and Irgarol are low, whereas those posed by DCOIT are high. However, the rate of detection of DCOIT was low because it degraded rapidly in the seawater before treatment for chemical analysis, except in the case of the treatment on the research vessel., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

In this study, we screened for the antifouling activity of 15 species plant extracts from Brazilian the Brazilian Caatinga Fabaceae against the initial colonization of natural marine bacterial biofilm. We also investigated the potential toxicity of extracts against planktonic and benthic non-target organisms. Aqueous extracts of plants collected in the Caatinga biome (PE, Brazil) were prepared and tested at different concentration levels (0, 0.5, 1, 2, 4, and 8 mg mL -1 ). Natural marine bacterial consortium was inoculated in multi-well plates and incubated with the different treatments for 48 h. The biofilm and planktonic bacterial density and biomass inhibition were evaluated along with biofilm biomass eradication. The extracts that showed the highest bacterial biofilm inhibition were evaluated for toxicity against microalgae and crustaceans. The biofilm and planktonic bacterial inhibition potential were evaluated through flow cytometry and spectrophotometry. The selected treatments were evaluated for their toxicity using the microalgae Chaetoceros calcitrans, the copepod Nitokra sp., and the brine shrimp Artemia salina as bioindicators. Our work demonstrates the biotechnological potential of Fabaceae plant compounds as a safe antifouling alternative. Anadenanthera colubrina var. cebil fruits and Apuleia leiocarpa leaf extracts showed antibiofilm activity (≥ 80%), while Myroxylon peruiferum and Dioclea grandiflora leaf extracts showed antibiotic activity. These extracts were safe to planktonic and benthic non-target organisms. The results of this study point to potential substitutes to highly toxic antifouling paints and shed light on the prospect of a yet to be explored biome for more sustainable alternatives in biofouling research.

A suitable model to predict the toxicity of current and continuously emerging disinfection by-products (DBPs) is needed. This study aims to establish a reliable model for predicting the cytotoxicity of DBPs to Chinese hamster ovary (CHO) cells. We collected the CHO cytotoxicity data of 74 DBPs as the endpoint to build linear quantitative structure-activity relationship (QSAR) models. The linear models were developed by using multiple linear regression (MLR). The MLR models showed high performance in both internal (leave-one-out cross-validation, leave-many-out cross-validation, and bootstrapping) and external validation, indicating their satisfactory goodness of fit (R 2  = 0.763-0.799), robustness (Q 2 LOO  = 0.718-0.745), and predictive ability (CCC = 0.806-0.848). The generated QSAR models showed comparable quality on both the training and validation levels. Williams plot verified that the obtained models had wide application domains and covered the 74 structurally diverse DBPs. The molecular descriptors used in the models provided comparable information that influences the CHO cytotoxicity of DBPs. In conclusion, the linear QSAR models can be used to predict the CHO cytotoxicity of DBPs.

We used a green sol-gel synthesis method to fabricate a novel nanoporous copper aluminosilicate (CAS) material. Nanoporous CAS was characterized using X-ray powder diffraction (XRD), field emission transmission and scanning electron microscopies (FE-TEM/FE-SEM), Fourier transform infrared (FTIR) spectroscopy, and optical analyses. The CAS was also evaluated for use as a promising disinfectant for the inactivation of waterborne pathogens. The antimicrobial action and minimum inhibitory concentration (MIC) of this CAS disinfectant were determined against eight microorganisms (Escherichia coli, Salmonella enterica, Pseudomonas aeruginosa, Listeria monocytogenes, Staphylococcus aureus, Enterococcus faecalis, Candida albicans, and Aspergillus niger). An antimicrobial susceptibility testing of CAS was measured. Results of disc diffusion method pointed out that the diameters of the zone using well diffusion were wider than disc diffusion methods, and the findings also showed that the MIC of the CAS disinfectant against E. coli, S. enterica, and P. aeruginosa was 100 mg/L within 20 min of contact time. Meanwhile, the MIC of the CAS disinfectant was 100 mg/L within 40 min of contact time for the other strains. The efficacy of antimicrobial action (100%) reached within 20 to 40 min against all tested microbes. Herein, the antimicrobial susceptibility testing of CAS disinfectant showed no toxicity for human and bacterial cells. It can be concluded that nanoporous CAS is a promising, economically, and worthy weapon for water disinfection.

The organic toxicants formed in chlorinated water cause potential harm to human beings, and it is extensively concentrated all over the world. Various disinfection by-products (DBPs) occur in chlorinated water are genotoxic and carcinogenic. The toxicity is major concern for chlorinated DBPs which has been present more in potable water. The purpose of the work was to evaluate genotoxic properties of DBPs in Allium cepa as a plant model system. The chromosomal aberration and DNA laddering assays were performed to examine the genotoxic effect of trichloroacetic acid (TCAA), trichloromethane (TCM), and tribromomethane (TBM) in a plant system with distinct concentrations, using ethyl methanesulfonate (EMS) as positive control and tap water as negative control. In Allium cepa root growth inhibition test, the inhibition was concentration dependent, and EC 50 values for trichloroacetic acid (TCAA), trichloromethane (TCM), and tribromomethane (TBM) were 100 mg/L, 160 mg/L, and 120 mg/L respectively. In the chromosome aberration assay, root tip cells were investigated after 120 h exposure. The bridge formation, sticky chromosomes, vagrant chromosomes, fragmented chromosome, c-anaphase, and multipolarity chromosomal aberrations were seen in anaphase-telophase cells. It was noticed that with enhanced concentrations of DBPs, the total chromosomal aberrations were more frequent. The DNA damage was analyzed in roots of Allium cepa exposed with DBPs (TCAA, TCM, TBM) by DNA laddering. The biochemical assays such as lipid peroxidation, H 2 O 2 content, ascorbate peroxidase, guaiacol peroxidase, and catalase were concentration dependent. The DNA interaction studies were performed to examine binding mode of TCAA, TCM, and TBM with DNAs. The DNA interaction was evaluated by spectrophotometric and spectrofluorometric studies which revealed that TCAA, TCM, and TBM might interact with Calf thymus DNA (CT- DNA) by non-traditional intercalation manner.

Ester-functionalized pyridinium ionic liquids (ILs), 1-decyloxycarbonylmethylpyridinium chloride (PyrСOOC 10 -Cl), and 1-dodecyloxycarbonylmethylpyridinium chloride (PyrСOOC 12 -Cl) have been synthesized and studied for their environmental toxicity. Simple long-chain pyridinium ILs, 1-dodecylpyridinium chloride (PyrC 12 -Cl), and commercial disinfectant cetylpyridinium chloride (CPC) were used as reference compounds. Both ester-functionalized ILs and CPC showed significantly reduced antibacterial activity compared to PyrC 12 -Cl. However, ester-functionalized ILs were found to have excellent antifungal activity towards Candida albicans fungus strains, similar to PyrC 12 -Cl and much higher than for CPC. The molecular docking of ILs in the active site of the known antifungal target N-myristoyltransferase (Nmt) C. albicans has been conducted. The obtained results indicate the possibility of ILs binding into the Nmt pocket. The high stability of the complexes, especially for PyrCOOC 10 -Cl, is ensured by hydrogen bonding, electrostatic anion-pi interactions, as well as hydrophobic pi-alkyl and alkyl interactions that was confirmed by calculated binding energy values. The acute toxicity studies of ester-functionalized ILs on D. rerio (zebrafish) hydrobiont have shown their dramatically reduced ecotoxicity compared to PyrC 12 -Cl and CPC. Thus, LD 50 values of 15.2 mg/L and 16.8 mg/L were obtained for PyrCOOC 10 -Cl and PyrCOOC 12 -Cl, respectively, whereas CPC had LD 50 value of 0.018 mg/L. The primary biodegradation test CEC L-33-A93 of ILs indicated an improved biodegradability of ester-functionalized compounds compared to simple long-chain ILs. Based on the obtained results, PyrCOOC 10 -Cl may be considered as very promising cationic biocide due to the combination of soft antimicrobial activity and reduced ecotoxicity, as well as improved biodegradability.

A combined use of silver nanoparticles (AgNPs) with different types of disinfectants as antimicrobial might be useful in mitigating the problem of development of bacterial resistance with a strong enhancement of the biocidal effect of disinfectants. To evaluate the biocidal activity of silver nanoparticles and its loaded forms, five commercial disinfectants (quaternary ammonium compounds (benzalkonium chloride (BC) and TH 4+ ), Virkon®S, sodium hypochlorite, and hydrogen peroxide (H 2 O 2 )) were used against Listeria monocytogenes (L. monocytogenes) isolates at different concentrations and exposure times to reveal intra-species variability and the percentage of resistance to antimicrobial agents used. Therefore, a total of 260 specimens from animal and human stool as well as environmental samples from dairy cattle farms were cultured for isolation of L. monocytogenes. Thereafter, bacterial isolates were identified using PCR. Silver nanoparticle was synthesized using chemical reduction. Both silver nanoparticles and its loaded forms were characterized by transmission electron microscopy (TEM). The sensitivity test of 60 strains of L. monocytogenes bacteria to AgNPs and its loaded forms was evaluated using broth macrodilution method. Virkon®S/AgNPs 2.0% exhibited the highest bactericidal effect (100%) against L. monocytogenes strains followed by H 2 O 2 /AgNPs 5.0% and TH 4+ /AgNPs 1.0% (90% each). Furthermore, the percentage of resistance of L. monocytogenes was 0.0% to both H 2 O 2 /AgNPs 5.0% and Virkon®S/AgNPs 2.0%. In conclusion, monitoring the main source of contamination with Listeria monocytogenes in dairy cattle farms is an essential factor to achieve an efficient control. Moreover, the use of the disinfectants, Virkon®S 2.0%, H 2 O 2 5.0%, and TH 4+ 1.0%, loaded on silver nanoparticles composite had the strong bactericidal effect against L. monocytogenes.

Harmful algal blooms (HABs) induced by Prorocentrum donghaiense occur frequently and cause a serious threat to the marine ecosystem. In this study, antialgal effects of α-linolenic acid (ALA) that is generally extracted from diverse macroalga on P. donghaiense were investigated. Specifically, the growth, cellular morphology and ultrastructure, reactive oxygen species (ROS) content, mitochondrial membrane potential (MMP), cytochrome C (Cyt-C), and caspase-9,3 activity of untreated and treated P. donghaiense were investigated. The results showed that ALA significantly inhibited the growth of P. donghaiense. Under ALA exposure, the cellular morphology and ultrastructure were damaged. ALA also induced ROS overproduction in the algal cells, decreased MMP, induced Cyt-C release, and activated caspase-9,3, which strongly relates to algal apoptosis. In summary, this study revealed the responses of morphology and physiology of P. donghaiense when exposed under ALA, and shows the potential of biotechnology on controlling P. donghaiense.

Antifouling chemicals are legacy contaminants in aquatic ecosystems. Previous experiments have shown that a 14-day exposure to the antifouling chemical medetomidine delays metamorphosis and reduces body mass in wood frog tadpoles. In the present study, we exposed wood frog tadpoles to medetomidine for 3, 7, and 10 days at 100 nM, 1 μM, and 10 μM. We also exposed American toad tadpoles to medetomidine for 3 days at four concentrations (10 nM, 100 nM, 1 μM, and 10 μM) in static renewal experiments. In each experiment, we measured growth, frequency and time to metamorphosis, and mass at metamorphosis. In both species, medetomidine significantly slowed development as measured by the Gosner stage. After 34 days in culture, wood frog tadpoles exposed to 1 and 10 μM medetomidine for as few as 3 days were significantly less developed compared to controls. Toads exposed to 1 μM medetomidine for 3 days were also significantly less developed on day 27, but by day 34, there was no difference from controls. For wood frogs, medetomidine significantly affected time to metamorphosis with a trend for tadpoles at lower concentrations metamorphosing sooner than those at higher concentrations. While medetomidine affected time to metamorphosis in wood frogs, it did not affect fresh mass, dry mass, or mortality compared to controls. Wood frog tadpoles that did not metamorphose after over 90 days in culture were more frequent in high-concentration groups than in the control. In toads, 10 μM medetomidine was 100% lethal within 23 days, but at the same concentration and duration, no wood frog tadpoles died. Lower concentrations were also significantly lethal to toads compared to controls, but tadpoles that survived in 10 and 100 nM metamorphosed sooner than those in 1 μM. Fresh mass of toad tadpoles exposed to 1 μm was significantly smaller at metamorphosis compared to that of controls. Medetomidine also affected the behavior of tadpoles. In toads, medetomidine significantly reduced both percent activity and startle response. In wood frogs, medetomidine significantly reduced percent activity, but increased startle response. We discuss our finding of low-dose stimulation and high-dose inhibition of different life history endpoints in terms of hormetic mechanisms. The differential sensitivity between species in terms of mortality, frequency of metamorphosis, and behavior highlights the potential negative environmental effects of medetomidine to amphibians.

Toxic effects of continuous low dose application of the antifouling biocide chlorine on marine benthic organisms were monitored using transplanted green mussels (Perna viridis) and a suite of biomarkers. Caged mussels were deployed in chlorinated and non-chlorinated sections of the cooling system of an operating electric power plant. Biomarkers indicative of general stress, oxidative stress (superoxide dismutase and catalase), and DNA integrity, along with expression of stress proteins, were studied to assess the effects. Deterioration in condition index with corresponding increase in DNA strand breaks was indicative of chlorine stress. Superoxide dismutase enzyme did not show any particular trend, but catalase activity was high during the initial days of exposure at the chlorinated site; later, it became almost equal to that at the control site. Similarly, expressions of stress proteins (HSP60, HSP70, HSP22, GSTS1, and CYP4) showed bell-shaped pattern during the period of study. Positive correlation among the endpoints indicated the utility of the multimarker approach to monitor the effects of continuous low dose chlorination on mussels.

Tributyltin (TBT) is a biocide extremely toxic to a wide range of organisms, which has been used for decades for industrial purposes. Fucoxanthin is a natural carotenoid that is isolated from seaweed, and fucoxanthinol is a major primary metabolite of fucoxanthin. Although fucoxanthin and fucoxanthinol have been reported to possess anti-oxidant activities in vitro, little is known as to whether they protect against TBT-induced oxidative stress in cultured cells. In the present study, the protective effect of fucoxanthin and fucoxanthinol against oxidative stress induced by TBT was investigated. The data showed that incubation of HepG2 cells with 0.2 μM TBT significantly increased cell apoptosis, whereas treatment with fucoxanthin or fucoxanthinol (3 μM) significantly recovered cell viability. In addition, fucoxanthinol treatment significantly decreased the intracellular reactive oxygen species (ROS) and malondialdehyde (MDA) in HepG2 cells incubated with TBT. Moreover, fucoxanthin and fucoxanthinol markedly increased the expression level of Bcl-2/Bax. These results demonstrated that both fucoxanthin and fucoxanthinol effectively prevented cytotoxicity in HepG2 cells treated with TBT, and the protective effect was likely associated with decreased intracellular ROS and MDA and increased Bcl-2/Bax levels.

This study was aimed to investigate the antifouling (AF) potentials and toxic properties of methanol extract from leaves of mangrove Excoecaria agallocha. Antimicrofouling activity results inferred that this extract strongly inhibited fouling bacterial and microalgal growth. This extract had also inhibited the settlement of brown mussel Perna indica and larvae of barnacle Balanus amphitrite. Further, EC 50 < LC 50 and therapeutic ratio > 1 together propagated non-toxic nature of the extract. Mollusk foot adherence assay result showed complete inhibition of foot spreading and loss of attachment of common rocky fouler Patella vulgata to the substrata. Field assay results affirmed that this extract effectively deterred settlement of biofoulers. Purification and GC-MS analysis of bioassay-guided active spot evidenced presence of three major compounds (> 85%) responsible for the promising AF activity. The identified lead compounds subjected to an estimation (BIOWIN™) program developed by United States Environmental Protection Agency (USEPA) predicts that they are biodegradable in nature. Graphical abstract.

Sodium chlorate (NaClO 3 ) is a widely used non-selective herbicide. It is also generated as a byproduct during disinfection of drinking water by chlorine dioxide. In the present work, the effects of NaClO 3 on human erythrocytes were studied under in vitro conditions. Incubation of erythrocytes with different concentrations of NaClO 3 at 37 °C for 90 min resulted in significant hemolysis. Cell lysates were prepared from NaClO 3 -treated and untreated (control) erythrocytes and assayed for various biochemical parameters. Methemoglobin levels were significantly increased and methemoglobin reductase activity was reduced upon NaClO 3 treatment. There was a significant increase in protein oxidation and lipid peroxidation with a decrease in reduced glutathione and total sulfhydryl content. This suggests the induction of oxidative stress in erythrocytes upon exposure to NaClO 3 . The occurrence of oxidative stress was confirmed by significantly increased generation of reactive oxygen species and lowered antioxidant response of the cells. NaClO 3 treatment also increased nitric oxide levels showing induction of nitrosative stress. The activities of major antioxidant and membrane-bound and metabolic enzymes were significantly altered upon incubation of erythrocytes with NaClO 3 . The erythrocytes became more osmotically fragile while electron microscopic images showed gross morphological alterations in NaClO 3 -treated cells. These results show that NaClO 3 induces oxidative stress in human erythrocytes, which results in extensive membrane damage and lowers the antioxidant response.

Plant-based bioassays have gained wide use among the toxicological and/or ecotoxicological assessment procedures because of their simplicity, sensitivity, low cost, and reliability. The present study describes the use of Vicia faba (V. faba) micronucleus (MN) test and V. faba comet assay in the evaluation of the genotoxic potential of disinfection by-products (DBPs) commonly found in chlorine-disinfected drinking water. Five haloacetic acids and three halogenated acetonitriles were chosen as representatives of DBPs in this study because they are of potentially great public health risk. Results of the MN test indicated that monochloroacetic acid (MCA), monobromoacetic acid (MBA), dichloroacetic acid (DCA), dibromoacetic acid (DBA), trichloroacetic acid (TCA), and trichloroacetonitrile (TCAN) caused a statistically significant increase in MN frequency in V. faba root tip cells. However, no genotoxic response was observed for dichloroacetonitrile (DCAN) and dibromoacetonitrile (DBAN). Results of the comet assay showed that all tested DBPs induced a statistically significant increase in genomic DNA damage to V. faba root tip cells. On considering the capacity to detect genomic damage of a different nature, we suggest that a combination of V. faba MN test and V. faba comet assay is a useful tool for the detection of genotoxic effects of DBPs. It is worthy of assessing the feasibility of using V. faba comet assay combined with V. faba MN test to screen for the genotoxic activity of chlorinated drinking water in future work.

The biocide Irgarol 1051 has been reported to have negative effects on a large number of living components including non-target organisms, but information on its impact on the marine meiofauna and benthic prokaryotes is completely lacking. Here, we report the results of long-term experimental studies in which we determined the effects of increasing Irgarol concentrations (from 11.5 to 315 ng g(-1) sediment dry weight) on meiofauna and benthic prokaryotes. We found that this biocide had a significant impact on meiofauna abundance, even at the lowest concentrations, causing a drastic decline in the abundance of nematodes (the dominant meiofaunal taxon) and an increase of the relative importance of oligochaetes. Even if no direct effects of Irgarol were found on prokaryotic abundance and biomass, the molecular fingerprinting analyses (automated ribosomal intergenic spacer analysis) showed that the prokaryotic diversity was significantly altered by the biocide. The results of the present study indicate that Irgarol 1051 in marine sediments has a significant impact on the smallest eukaryotic and microbial components also at very low concentrations (ca 12 ng g(-1)).

We tested the effects of solvent fractions (FA, FB, FC, and FD), which partitioned by liquid-liquid extraction from the methanol extract of Ulva pertusa, on the growth of red tide microalgae (Karenia mikimitoi, Skeletonema costatum, Alexandrium tamarense, Heterosigma akashiwo, Prorocentrum donghaiense), and FA, FB, and FC exhibited significantly antialgal activity. The chemical constituent analysis showed the existence of bioactive compounds such as phenols and alkaloids. Further, four solvent fractions were applied to silica gel column and repeated preparative TLC to produce 13 samples and their purity qualified as thin-layer chromatographic grade. Among these purified samples, FA111, FB411, FC411, FD111, and FD211 exhibited stronger antialgal activity. Furthermore, their functional groups were analyzed by colorimetric methods and UV spectra data. FD111 and FD211 were temptatively identified as alkaloids; the others were initially identified as phenolic acids. This is a preliminary study and the structure identification of these purified samples requires further investigation. While concentration of these purified samples in this algae was very small, they showed excellent effects against red tide microalgae.

A 2-year study was implemented to characterize the contamination of estuarine continuums in the Bay of Vilaine area (NW Atlantic Coast, Southern Brittany, France) by 30 pesticide and biocide active substances and metabolites. Among these, 11 triazines (ametryn, atrazine, desethylatrazine, desethylterbuthylazine, desisopropyl atrazine, Irgarol 1051, prometryn, propazine, simazine, terbuthylazine, and terbutryn), 10 phenylureas (chlortoluron, diuron, 1-(3,4-dichlorophenyl)-3-methylurea, fenuron, isoproturon, 1-(4-isopropylphenyl)-3-methylurea, 1-(4-isopropylphenyl)-urea, linuron, metoxuron, and monuron), and 4 chloroacetanilides (acetochlor, alachlor, metolachlor, and metazachlor) were detected at least once. The objectives were to assess the corresponding risk for aquatic primary producers and to provide exposure information for connected studies on the responses of biological parameters in invertebrate sentinel species. The risk associated with contaminants was assessed using risk quotients based on the comparison of measured concentrations with original species sensitivity distribution-derived hazardous concentration values. For EU Water Framework Directive priority substances, results of monitoring were also compared with regulatory Environmental Quality Standards. The highest residue concentrations and risks for primary producers were recorded for diuron and Irgarol 1051 in Arzal reservoir, close to a marina. Diuron was present during almost the all survey periods, whereas Irgarol 1051 exhibited a clear seasonal pattern, with highest concentrations recorded in June and July. These results suggest that the use of antifouling biocides is responsible for a major part of the contamination of the lower part of the Vilaine River course for Irgarol 1051. For diuron, agricultural sources may also be involved. The presence of isoproturon and chloroacetanilide herbicides on some dates indicated a significant contribution of the use of plant protection products in agriculture to the contamination of Vilaine River. Concentration levels and associated risk were always lower in estuarine sites than in the reservoir, suggesting that Arzal dam reduces downstream transfer of contaminants and favors their degradation in the freshwater part of the estuary. Results of the additional monitoring of two tidal streams located downstream of Arzal dam suggested that, although some compounds may be transferred to the estuary, their impact was probably very low. Dilution by marine water associated with tidal current was also a major factor of concentration reduction. It is concluded that the highest risks associated to herbicides and booster biocides concerned the freshwater part of the estuary and that its brackish/saltwater part was exposed to a moderate risk, although some substances may sometimes exhibit high concentration but mainly at low tide and on an irregular basis.

Background, Aim, and Scope: Glutaraldehyde (GA) often acts as an effective sterilant, disinfectant, and preservative in chemical products. It was found that GA had clearly acute toxicity to aquatic organisms. Furthermore, GA in natural environment could not exist as single species but as complex mixtures. To explore the toxicity interaction between GA and the other environmental pollutant, it is necessary to determine the mixture toxicities of various binary mixtures including GA. Two reference models, concentration addition (CA) and independent action (IA), are often employed to evaluate the mixture toxicity, which can be finished by comparing the concentration-response curves (CRCs) predicted by the reference models with the experimental CRC of the mixture. However, the CRC-based method cannot effectively denote the degree of the deviations from the reference models, especially at very low effect levels. Though the model deviation ratio (MDR) can be used to quantitatively evaluate the deviation of a mixture at EC50 level from the reference model, it is difficult to evaluate the deviations at the lower effect levels. Therefore, the primary aim of this study was to develop a new effect residual ratio (ERR) method to validate the deviations from the reference models at various effect levels., Materials and Methods: Four chemicals having possible dissimilar mode of actions with GA, acetonitrile (ACN), dodine (DOD), simetryn (SIM), and metham sodium (MET), were selected as another component in the binary mixtures including GA, which constructed four binary mixtures, GA-ACN, GA-DOD, GA-SIM, and GA-MET ones. For each binary mixture, two equipotent mixture rays where the concentration ratios of GA to another mixture component are respectively EC50 and EC5 ones were designed and their toxicities (expressed as a percent inhibition to Photobacterium phosphoreum) were determined by microplate toxicity analysis. The observed concentration-response curve (CRC) of a ray was compared with that predicted by CA or IA model to qualitatively assess the toxicity interaction of the mixture ray. To quantitatively and effectively examine the deviations at various effect levels from the reference models, a new concept, ERR at an effect, was defined, and the ERR was employed to evaluate the deviation at various effects with confidence intervals., Results: For three binary mixtures, GA-ACN, GA-DOD, and GA-SIM, the CRCs predicted by IA models were almost located in the 95% confidence intervals of the experimental CRCs for both equipotent mixture rays, which indicated the independent actions between binary mixture components. However, two rays of GA-MET binary mixture displayed a little synergistic action because both CRCs predicted by CA and IA were lower than the experimental CRC. ERR showed the same results as MDR, but ERR results at low effect area were clearer than MDR ones., Discussion: In CRC comparison, the deviation of CA (for GA-ACN, GA-DOD, and GA-SIM combinations) or IA (for GA-MET) model from the experimental values could be obviously observed at medium area of the CRC. However, at very low effect levels, both deviations of CA and IA and difference between CA and IA model predictions were not very apparent. Thus, it was difficult to confirm which model, CA or IA, had better predicted power at very low effect levels. MDR in many literatures often refers to a ratio at EC50 level. It was also difficult to reflect not only the deviation fact at the other ECx but also the deviation uncertainty. After we extended the definition of MDR to all ECx and examined the 95% confidence intervals based on observation, the plot of the redefined MDRs at many effect levels could better explain the deviations of CA or IA model from the observation. However, MDRs at very low effect levels did not still reflect the high uncertainty there. The ERRs defined in our paper could explicitly explain the degree of deviation from the reference models and especially reflect the high uncertainty at very low effects. It could be said that the ERR is a better indicator than MDR., Conclusions: The new ERR validation method developed in our laboratory could provide us with the information about the toxicity interaction between the mixture components and quantitatively assess the accuracy of the reference models (CA or IA) at whole effect levels. The ERR method conquered the invalidation of the classical CRC comparison method on the deviation decision at low effect levels and also got the advantage over the MDR methods., Recommendations and Perspectives: It holds promise to become an effective method of hazard and risk assessments of chemical mixtures by well characterizing the uncertainty at very low effect levels.