Multi- and transgenerational effects of environmental chemicals on mollusks: An underexplored experimental design in aquatic (eco)toxicological studies.

(Eco)toxicological studies frequently evaluate the effects of chemicals in one life stage of organisms, but the use of these outcomes can only partially estimate populational effects. In this regard, multi- and/or transgenerational studies should be performed in order to provide information on contaminant effects in a populational functioning context. The present review aimed to summarize and critically evaluate the current knowledge regarding multi- and/or transgenerational effects of traditional and emerging environmental chemicals on mollusks. Results showed that these kinds of studies were performed in aquatic mollusks (bivalve and gastropod), being Gastropoda the mollusk Class most frequently studied. Additionally, freshwater species and multigenerational studies were more common for this class. For the Bivalvia class, only marine species were evaluated, and transgenerational exposure was more commonly assessed. The effects were reported for 15 species, highlighting the marine bivalves Crassostrea gigas and Saccostrea glomerata, and the freshwater gastropod Lymnaea stagnalis. Multi- and transgenerational effects were described for 8 environmental chemical groups, mainly metals, pesticides, and pharmaceuticals. In general, multi- and transgenerational exposure induced biometric, developmental, and reproductive impairments in mollusks, indicating that environmental chemicals might lead to generational impairments, reduced population growth and reproductive capacity, and decreased fitness. The current study indicated that bivalves and gastropods are suitable organism models to assess the multi- and transgenerational adverse effects induced by traditional and emerging environmental chemicals.

Green synthesis of silver nanoparticles using Croton urucurana and their toxicity in freshwater snail species Biomphalaria glabrata.

Green silver nanoparticles (G-Ag NPs) have contributed to the development of ecological technologies with low environmental impact and safer for human health, as well as demonstrating potential for the control of vectors and intermediate hosts. However, knowledge about its toxicity in the early stages of gastropod development remains scarce. Therefore, the current study aimed to investigate the toxicity of G-Ag NPs synthesized from Croton urucurana leaf extracts in snail species Biomphalaria glabrata, which is an intermediate host for Schistosoma mansoni parasite. G-Ag NPs were synthesized using two types of plant extracts (aqueous and hydroethanolic) and characterized using multiple techniques. Bioassays focused on investigating G-Ag NPs and plant extracts were carried out with embryos and newly hatched snails, for 144 h and 96 h, respectively; toxicity was analyzed based on mortality, hatching, development inhibition, and morphological changes. Results have shown that both G-Ag NPs were more toxic to embryos and newly hatched snails than the investigated plant extracts. G-Ag NPs deriving from aqueous extract have higher molluscicidal activity than those deriving from hydroethanolic extract. Both G-Ag NPs induced mortality, hatching delay, development inhibition, and morphological changes (i.e., hydropic embryos), indicating their molluscicidal activities. Moreover, embryos were more sensitive to G-Ag NPs than newly hatched snails. Thus, the toxicity of G-Ag NPs to freshwater snails depends on the type of extracts and the snail's developmental stages. These findings can contribute to the development of green nanobiotechnologies applicable to control snails of medical importance.

Toxicity Assessment of New Ag-ZnO/AgO Nanocomposites: An In Vitro and In Vivo Approach.

Zinc oxide nanoparticles (ZnO NPs) are metal oxide nanomaterials, which are important for several applications: antibacterial, anthelmintic, antiprotozoal and antitumoral, among others. These applications are mainly related to the ability to spontaneously produce and induce the production of reactive oxygen species that are important components for the destruction of pathogens and tumor cells. While trying to potentiate ZnO NPs, studies have associated these NPs with silver oxide (AgO) or silver (Ag) NPs. It has already been reported that this combination (Ag-ZnO/AgO NPs) is able to enhance the microbicidal potential. Although possessing much potential for several purposes, it is important to evaluate whether this association also poses the risk of toxicity to cells and experimental models. Therefore, this work aimed to evaluate the toxicity of various Ag-ZnO/AgO NP nanocomposites, in vitro and in vivo. Accordingly, ZnO nanocrystals and nanocomposites with various concentrations of AgO (ZnO:5Ag, ZnO:9Ag or ZnO:11Ag) were used in different cytotoxicity models: Galleria mellonella (G. mellonella), cell lines (VERO and RAW 264.7) and C57BL/6 mice. In the G. mellonella model, four concentrations were used in a single dose, with subsequent evaluation of mortality. In the case of cells, serial concentrations starting at 125 µg/mL were used, with subsequent cytotoxicity assessment. Based on the safe doses obtained in G. mellonella and cell models, the best doses were used in mice, with subsequent evaluations of weight, biochemistry as also renal and liver histopathology. It was observed that the toxicity, although low, of the nanocomposites was dependent upon the concentration of AgO used in association with ZnO NPs, both in vitro and in vivo.

Microplastics and TiO2 nanoparticles mixture as an emerging threat to amphibians: A case study on bullfrog embryos.

Emerging contaminants can act as contributing factors to the decline of amphibian populations worldwide. Recently, scientists have drawn attention to the potential ecotoxicity of microplastics and nanomaterials in amphibians, however, their possible effects on embryonic developmental stages are still absent. Thus, the present study analyzed the developmental toxicity of environmentally relevant concentrations of polyethylene microplastics (PE MPs; 60 mg/L) and titanium dioxide nanoparticles (TiO2 NPs; 10 µg/L), isolated or in combination (Mix group) on bullfrog embryos, Aquarana catesbeiana, adapting the Frog Embryo Teratogenesis Assay (FETAX, 96h). Allied to the FETAX protocol, we also analyzed the heart rate and morphometric data. The exposure reduced the survival and hatching rates in groups exposed to TiO2 NPs, and to a lesser extent, also affected the Mix group. TiO2 NPs possibly interacted with the hatching enzymes of the embryos, preventing hatching, and reducing their survival. The reduced effects in the Mix group are due to the agglomeration of both toxicants, making the NPs less available for the embryos. PE MPs got attached to the gelatinous capsule of the chorion (confirmed by fluorescence microscopy), which protected the embryos from eventual direct effects of the microplastics on the hatching and survival rates. Although there were no cardiotoxic effects nor morphometric alterations, there was a significant increase in abdominal edemas in the hatched embryos of the PE MPs group, which indicates that osmoregulation might have been affected by the attachment of the microplastics on the embryos' gelatinous capsule. This study presents the first evidence of developmental toxicity of environmental mixtures of microplastics and nanoparticles on amphibians and reinforces the need for more studies with other amphibian species, especially neotropical specimens that could present bigger sensibility. Our study also highlighted several features of the FETAX protocol as useful tools to evaluate the embryotoxicity of several pollutants on amphibians.

Zebrafish neuromast sensory system: Is it an emerging target to assess environmental pollution impacts?

Environmental pollution poses risks to ecosystems. Among these risks, one finds neurotoxicity and damage to the lateral line structures of fish, such as the neuromast and its hair cells. Zebrafish (Danio rerio) is recommended as model species to be used in ecotoxicological studies and environmental biomonitoring programs aimed at assessing several biomarkers, such as ototoxicity. However, little is known about the history of and knowledge gaps on zebrafish ototoxicity. Thus, the aim of the current study is to review data available in the scientific literature about using zebrafish as animal model to assess neuromast toxicity. It must be done by analyzing the history and publication category, world production, experimental design, developmental stages, chemical classes, neuromasts and hair cell visualization methods, and zebrafish strains. Based on the results, number, survival and fluorescence intensity of neuromasts, and their hair cells, were the parameters oftentimes used to assess ototoxicity in zebrafish. The wild AB strain was the most used one, and it was followed by Tübingen and transgenic strains with GFP markers. DASPEI was the fluorescent dye most often applied as method to visualize neuromasts, and it was followed by Yo-Pro-1 and GFP transgenic lines. Antibiotics, antitumorals, metals, nanoparticles and plant extracts were the most frequent classes of chemicals used in the analyzed studies. Overall, pollutants can harm zebrafish's mechanosensory system, as well as affect their behavior and survival. Results have shown that zebrafish is a suitable model system to assess ototoxicity induced by environmental pollution.

2,4-D Herbicide-Induced Hepatotoxicity: Unveiling Disrupted Liver Functions and Associated Biomarkers.

2,4-dichlorophenoxyacetic acid (2,4-D) is a widely used herbicide worldwide and is frequently found in water samples. This knowledge has prompted studies on its effects on non-target organisms, revealing significant alterations to liver structure and function. In this review, we evaluated the literature on the hepatotoxicity of 2,4-D, focusing on morphological damages, toxicity biomarkers and affected liver functions. Searches were conducted on PubMed, Web of Science and Scopus and 83 articles were selected after curation. Among these studies, 72% used in vivo models and 30% used in vitro models. Additionally, 48% used the active ingredient, and 35% used commercial formulations in exposure experiments. The most affected biomarkers were related to a decrease in antioxidant capacity through alterations in the activities of catalase, superoxide dismutase and the levels of malondialdehyde. Changes in energy metabolism, lipids, liver function, and xenobiotic metabolism were also identified. Furthermore, studies about the effects of 2,4-D in mixtures with other pesticides were found, as well as hepatoprotection trials. The reviewed data indicate the essential role of reduction in antioxidant capacity and oxidative stress in 2,4-D-induced hepatotoxicity. However, the mechanism of action of the herbicide is still not fully understood and further research in this area is necessary.

Polystyrene nanoplastics induce developmental impairments and vasotoxicity in zebrafish (Danio rerio).

The exponential use of plastics has significantly increased environmental pollution by nanoplastics (NPs). In the aquatic environment, NPs interact and bioaccumulate in the biota, posing a potential ecotoxicological risk. The present study investigated the developmental toxicity, vasotoxicity, cytotoxicity, ROS induction, and behavioral impairments in zebrafish (Danio rerio) exposed to environmentally relevant polystyrene NPs (PS-NPs) concentrations (0.04, 34 ng L-1, and 34 µg L-1) for 144 h through multiple biomarkers response (mortality, frequency of spontaneous contractions, heart rate, and morphological changes). Furthermore, vasotoxicity (head, yolk sac, tail, and branchial vessels) was evaluated using the transgenic zebrafish tg(Fli1:eGFP). Results showed that PS-NPs interacted mainly with zebrafish chorion, gills, tail, and larvae head. PS-NPs at 34 ng L-1 and 34 µg L-1 induced neurotoxicity (decreased frequency of spontaneous contractions), cardiotoxicity (bradycardia), and morphological changes in the eyes and head, indicating that PS-NPs induce developmental impairments in zebrafish. In addition, cytotoxicity in the caudal region (34 ng L-1), ROS production, decreased mean swimming speed, and distance covered were observed in all tested concentrations. PS-NPs also induced vasotoxicity (yolk sac region) in transgenic zebrafish. Overall, the present study demonstrates the harmful effects of PS-NPs on the early developmental stages of freshwater fish, indicating their environmental risk.

Exposure to microcrystallized cellulose affects the health of tadpoles and sheds light on the threat these materials pose to amphibians.

The increasing use of cellulose-based materials (CBMs) has provided beneficial applications in different sectors. However, its release into environments may represent an ecological risk, therefore demanding that ecotoxicological studies be conducted to understand the risks (current and future) of CBM pollution. Thus, we evaluated the possible effects of microcrystalline cellulose (CMs) in Physalaemus cuvieri tadpoles. After seven days of exposure to CMs (at 58.29 and 100 mg/L), the animals were subjected to behavioral evaluation, and different biomarkers (biometric and biochemical) were evaluated. Although our data do not point to a neurotoxic effect of CMs (inferred by the absence of behavioral changes and changes in AChE and BChE activity), animals exposed to CMs showed differences in body condition. Furthermore, we noticed an increase in the frequency of erythrocyte nuclear abnormalities and DNA damage, which were correlated with the ingestion of CMs. We noticed that the antioxidant activity of tadpoles exposed to CMs (inferred by SOD, CAT, and DPPH radical scavenging activity) was insufficient to control the increase in ROS and MDA production. Furthermore, exposure to CMs induced a predominant Th2-specific immune response, marked by suppressed IFN-γ and increased IL-10 levels, with a consequent reduction in NO levels. Principal component analysis and IBRv-2 indicate, in general, a primarily more toxic response to animals exposed to the highest CM concentration. Therefore, our study evidence that CMs affect the health of P. cuvieri tadpoles and sheds light on the threat these materials pose to amphibians.

Carbon nanotubes and nanofibers seen as emerging threat to fish: Historical review and trends.

Since the discovery of the third allotropic carbon form, carbon-based one-dimensional nanomaterials (1D-CNMs) became an attractive and new technology with different applications that range from electronics to biomedical and environmental technologies. Despite their broad application, data on environmental risks remain limited. Fish are widely used in ecotoxicological studies and biomonitoring programs. Thus, the aim of the current study was to summarize and critically analyze the literature focused on investigating the bioaccumulation and ecotoxicological impacts of 1D-CNMs (carbon nanotubes and nanofibers) on different fish species. In total, 93 articles were summarized and analyzed by taking into consideration the following aspects: bioaccumulation, trophic transfer, genotoxicity, mutagenicity, organ-specific toxicity, oxidative stress, neurotoxicity and behavioral changes. Results have evidenced that the analyzed studies were mainly carried out with multi-walled carbon nanotubes, which were followed by single-walled nanotubes and nanofibers. Zebrafish (Danio rerio) was the main fish species used as model system. CNMs' ecotoxicity in fish depends on their physicochemical features, functionalization, experimental design (e.g. exposure time, concentration, exposure type), as well as on fish species and developmental stage. CNMs' action mechanism and toxicity in fish are associated with oxidative stress, genotoxicity, hepatotoxicity and cardiotoxicity. Overall, fish are a suitable model system to assess the ecotoxicity of, and the environmental risk posed by, CNMs.

The amphibian's spleen as a source of biomarkers for ecotoxicity assessment: Historical review and trends.

Amphibians are very sensitive to many environmental changes, so these animals are considered good bioindicator models for ecotoxicology. Given the importance of the amphibian spleen for hematopoietic and immune responses, this can be a key organ for the evaluation of biomarkers to monitor the health of individuals in nature or in captivity. In this systematic review, we searched databases and summarized the main findings concerning the amphibian spleen as a source of possible biomarkers applied in different scientific fields. The searches resulted in 83 articles published from 1923 to 2022, which applied the use of splenic samples to evaluate the effects of distinct stressors on amphibians. Articles were distributed in more than twenty countries, with USA, Europe, and Brazil, standing out among them. Publications focused mainly on anatomical and histomorphological characterization of the spleen, its physiology, and development. Recently, the use of splenic biomarkers in pathology and ecotoxicology began to grow but many gaps still need to be addressed in herpetological research. About 85 % of the splenic biomarkers showed responses to various stressors, which indicates that the spleen can provide numerous biomarkers to be used in many study fields. The limited amount of information on morphological description and splenic anatomy in amphibians may be a contributing factor to the underestimated use of splenic biomarkers in herpetological research around the world. We hope that this unprecedented review can instigate researchers to refine herpetological experimentation, using the spleen as a versatile and alternative source for biomarkers in ecotoxicology.

Cellulose microcrystalline: A promising ecofriendly approach to control Culex quinquefasciatus larvae.

The growing use of synthetic chemical compounds/substances in vector control of mosquitoes, associated with their adverse effects on the environment and non-target organisms, has demanded the development of eco-friendly alternatives. In this context, this study aimed to evaluate the larvicidal action of different cellulose microcrystalline (CMs) concentrations and investigate their toxicity mechanisms in Culex quinquefasciatus fourth instar larvae as a model species. Probit analysis revealed that the median lethal concentrations (LC50) for 24 h and 36 h exposure were 100 and 58.29 mg/L, respectively. We also showed that such concentrations induced a redox imbalance in the larvae, marked by an increase in the production of reactive oxygen species (ROS) and thiobarbituric acid reactive substances (TBARS), as well as a reduction in the activity of superoxide dismutase (SOD) and catalase (CAT). Furthermore, different alterations in the external morphology of the larvae were associated with the ingestion of CMs. On the other hand, exposure of adult zebrafish (Danio rerio) to LC5024h and LC5036h for seven days did not induce any behavioral changes or alterations mutagenic, genotoxic, biochemical, or in the production of cytokines IFN-γ and IL-10. Thus, taken together, our study demonstrates for the first time that the use of CMs can constitute a promising strategy in the control of C. quinquefasciatus larvae, combining insecticidal efficiency with an "eco-friendly" approach in the fight against an important mosquito vector of several human diseases.

A review on the ecotoxicity of macrocyclic lactones and benzimidazoles on aquatic organisms.

Despite its wide production and several applications, veterinary antiparasitics from macrocyclic lactones and benzimidazole classes have not received much scientific attention concerning their environmental risks. Thus, we aimed to provide insights into the state of the environmental research on macrocyclic lactone and benzimidazole parasiticides, emphasizing their toxicity to non-target aquatic organisms. We searched for relevant information on these pharmaceutical classes on PubMed and Web of Science. Our search yielded a total of 45 research articles. Most articles corresponded to toxicity testing (n = 29), followed by environmental fate (n = 14) and other issues (n = 2) of selected parasiticides. Macrocyclic lactones were the most studied chemical group (65% of studies). Studies were conducted mainly with invertebrate taxa (70%), with crustaceans being the most predominant group (n = 27; 51%). Daphnia magna was the most used species (n = 8; 15%). Besides, it also proved to be the most sensitive organism, yielding the lowest toxicity measure (EC50 0.25 µg/L for decreased mobility after 48 h-abamectin exposure) reported. Moreover, most studies were performed in laboratory settings, tracking a limited number of endpoints (acute mortality, immobility, and community disturbance). We posit that macrocyclic lactones and benzimidazoles warrant coordinated action to understand their environmental risks.

Plastic microfibers as a risk factor for the health of aquatic organisms: A bibliometric and systematic review of plastic pandemic.

Plastic microfibers (PMFs) are emerging pollutants widely distributed in the environment. In the early 2020s, the need for personal protection due to the COVID-19 pandemic led to increased consumption of plastic materials (e.g., facemasks and gloves) and ultimately to increased plastic pollution, especially by PMFs. The PMFs present in the environment may be released in this form (primary particles) or in larger materials, that will release them as a result of environmental conditions. Although a considerable number of studies have been addressing the effects of microplastics, most of them studied round particles, with fewer studies focusing on PMFs. Thus, the current study aimed to summarize and critically discuss the available data concerning the ecotoxicological impact of PMFs on aquatic organisms. Aquatic organisms exposed to PMFs showed accumulation, mainly in the digestive tract, and several toxic effects, such as DNA damage, physiological alterations, digestive damage and even mortality, suggesting that PMFs can pose a risk for the health of aquatic organisms. The PMFs induced toxicity to aquatic invertebrate and vertebrate organisms depends on size, shape, chemical association and composition of fibers. Regarding other size range (nm) of plastic fibers, the literature review highlighted a knowledge gap in terms of the effects of plastic nanofibers on aquatic organisms. There is a knowledge gap in terms of the interaction and modes of action of PMFs associated with other pollutants. In addition, studies addressing effects at different trophic levels as well as the use of other biological models should be considered. Overall, research gaps and recommendations for future research and trends considering the environmental impact of the COVID-19 pandemic are presented.

Bioaccumulation and ecotoxicological impact of micro(nano)plastics in aquatic and land snails: Historical review, current research and emerging trends.

Microplastics (MPs) and nanoplastics (NPs) are ubiquitous emerging pollutants in the environment. Although MPs/NPs' hazardous effects have been described at different trophic levels, little attention has been given to how they can affect gastropod communities. Thus, the current study aimed to summarize and critically address data available in the scientific literature about micro(nano)plastics' ecotoxicological impact on snails. The analyzed data has evidenced MP/NP bioaccumulation in 40 gastropod species collected in the field; 15 gastropod species were used to assess the potential toxicity of MPs/NPs. Asia accounted for the highest level of MPs/NPs bioaccumulated in gastropods; it was followed by the South American, European and Antarctic continents. MPs/NPs' toxicity depends on their composition, shape and size, as well as on differences in methodological approaches adopted by different studies. Results have shown that MPs/NPs induce several impairments - such as behavioral changes, developmental toxicity, dysbiosis, histopathological alterations, oxidative stress -, generate ecological impairments, as well as act as pollutant vector and increase chiral chemicals' toxicity. Research gaps and recommendations for future research were highlighted to help better understanding MPs/NPs' toxicity in gastropods, given the extremely important role played by them in studies focused on investigating how MPs/NPs can affect invertebrate communities living in terrestrial and aquatic environments.

Assessment of multiple biomarkers in Lithobates catesbeianus (Anura: Ranidae) tadpoles exposed to zinc oxide nanoparticles and zinc chloride: integrating morphological and behavioral approaches to ecotoxicology.

The ecotoxicological risk to vertebrates posed by zinc oxide nanoparticles (ZnO NPs) is still poorly understood, especially in animals with a biphasic life cycle, which have aquatic and terrestrial phases, such as amphibians. In the present study, we investigated whether acute exposure (7 days) to ZnO NPs and zinc chloride (ZnCl2) at three environmentally relevant concentrations (0.1, 1.0, and 10 mg L-1) induces changes in the morphology, chondrocranium, and behavior of the tadpoles of Lithobates catesbeianus (Anura: Ranidae). Tadpoles exposed to both forms of Zn did not undergo any morphological or behavioral changes at the lowest concentrations (0.1 and 1.0 mg L-1). However, the animals exposed to the highest concentration (10 mg L-1) lacked oral disc structures, were smaller in size, had a longer tail, and presented changes in the position and coiling of the intestine and malformations of the chondrocranium in comparison with the control group. This indicates that ZnO NPs and ZnCl2 altered the development of the tadpoles, causing delays in their metamorphosis and even reducing individual fitness. The tadpoles exposed to both forms of Zn at 10 mg L-1 also had reduced mobility, especially in the presence of conspecifics. Based on these findings, we emphasize the importance of studying morphological, skeletal, and behavioral biomarkers to evaluate the toxic effects of metal-based nanoparticles in amphibians.

Transgenic zebrafish (Danio rerio) as an emerging model system in ecotoxicology and toxicology: Historical review, recent advances, and trends.

Zebrafish (Danio rerio) is an alternative model system for drug screening, developing new products, and assessing ecotoxic effects of pollutants and biomonitor species in environmental risk assessment. However, the history and current use of transgenic zebrafish lines in ecotoxicology and toxicology studies remain poorly explored. Thus, the present study aimed to summarize and discuss the existing data in the literature about the applications of transgenic zebrafish lines in ecotoxicology and toxicology. The articles were analyzed according to publication year, journal, geographic distribution, and collaborations. Also, the bioassays were evaluated according to the tested chemical, transgenic lines, development stage, biomarkers, and exposure conditions (i.e., concentration, time, type, and route of exposure). Revised data showed that constitutive transgenic lines are the main type of transgenic used in the studies, besides most of uses embryos and larvae under static conditions. Tg(fli1: EGFP) was the main transgenic line, while the GFP and EGFP were the main reporter proteins. Transgenic zebrafish stands out in assessing vasotoxicity, neurotoxicity, systemic toxicity, hepatoxicity, endocrine disruption, cardiotoxicity, immunotoxicity, hematotoxicity, ototoxicity, and pancreotoxicity. This review showed that transgenic zebrafish lines are emerging as a suitable in vivo model system for assessing the mechanism of action and toxicity of chemicals and new biotechnology products, and the effects of traditional and emerging pollutants.

Toxicity of plant-based silver nanoparticles to vectors and intermediate hosts: Historical review and trends.

Green nanoparticles (GNPs), mainly green silver nanoparticles (Ag NPs), have been recommended as sustainable and eco-friendly technologies to control vectors and intermediate hosts. The aim of the current study is to carry out a historical and systematic literature review about the use of green plant-based Ag NPs (GP-Ag NPs) to control medically important mosquito, tick and gastropods. Data about the number of studies published per year, geographical distribution of studies (mailing address of the corresponding author), synthesis type (plant species, plant structure and extract types), physicochemical properties of GP-Ag NPs, experimental designs, developmental stages and the toxic effects on mosquitoes, ticks and gastropods were summarized and discussed. Revised data showed that GP-Ag NPs synthesis and toxicity in mosquitoes, ticks and snails depend on plant species, plant part, extract types, exposure condition and on the analyzed species. GP-Ag NPs induced mortality, tissue damage, biochemical and behavioral changes in mosquitoes and reduced their fecundity, oviposition, egg hatching and longevity. Ticks exposed to GP-Ag NPs presented increased mortality and reduced oviposition, while on snails, studies demonstrated mortality, oxidative stress, and DNA damage. Immune responses were also observed in snails after their exposure to GP-Ag NPs. GP-Ag NPs reduced the reproduction and population of several vectors and intermediate hosts. This finding confirms their potential to be used in gastropod control programs. Future studies about current gaps in knowledge are recommended.

Biomphalaria embryotoxicity test (BET): 60 years of research crossing boundaries for developing standard protocols.

Snail's embryotoxicity test is a suitable approach for toxicity assay of traditional and emerging pollutants, environmental risk assessment, as well as screening and development of new molluscicides. Among the snail species, Biomphalaria spp. has been indicated as a promising model system for developing standardized test protocols for assessing the chemical toxicity using early developmental stages. Thus, the current study aimed to review the data available in the scientific literature concerning the experimental approach, type of chemicals and the response of multiple biomarkers (survival, hatching rate, development delays, morphological and behavior changes) in snail embryos applied in toxicity tests. Revised data showed that the use of Biomphalaria embryos to assess chemical toxicity began in 1962. Snail's embryotoxicity test was applied mainly for analyzing the toxicity and development of new molluscicides, while its use in ecotoxicological studies is emerging. Biomphalaria glabrata was the main species analyzed. Embryos exposed to chemicals showed bioaccumulation, mortality, hatching inhibition, development delays, and morphological malformations, which were classified into four categories (hydropic, shell, cephalic and unspecified malformations). Besides, research gaps and recommendations for future research are indicated. Overall, the results showed that the Biomphalaria embryotoxicity test (BET) is a suitable tool for toxicity and health risk assessment.

Titanium dioxide nanoparticles as a risk factor for the health of Neotropical tadpoles: a case study of Dendropsophus minutus (Anura: Hylidae).

The production and use of titanium dioxide (TiO2) nanoparticles are increasing worldwide. The release of this substance into the environment can induce toxic effects in aquatic invertebrates and vertebrates, although the exact nature of its impacts on Neotropical amphibians is still poorly understood. In this context, the present study evaluated the toxicity of TiO2 nanoparticles and their counterpart-dissolved titanium dioxide (TiO2)-in the tadpoles of Dendropsophus minutus. The biometric parameters, DNA damage, and behavioral changes were verified in tadpoles exposed to three different concentrations (0.1 mg·L-1, 1.0 mg·L-1, and 10 mg·L-1) of TiO2 nanoparticles and dissolved TiO2 for 7 days. We verified significant DNA damage in the D. minutus tadpoles exposed to both forms of Ti, in comparison with the control group. We also identified a reduction in total size, body length, and width, and the height of the musculature of the tail of the tadpoles exposed to all concentrations of both substances in comparison with the control. In the behavioral test, the tadpoles exposed to nanoparticles and dissolved TiO2 presented reduced mobility and a tendency to be less aggregated than normal. Here, the simultaneous use of multiple biomarkers was fundamental for the reliable assessment of the adverse effects of nanomaterials on anuran amphibians and the establishment of a systematic approach to the biomonitoring of aquatic ecosystems. The present study expands our understanding of the genotoxic, morphological, and behavioral effects of TiO2 nanoparticles and dissolved TiO2 on anuran amphibians, and contributes to the establishment of further research for the more systematic assessment of the environmental risk of nanomaterials.

Toxicity evaluation of the combination of emerging pollutants with polyethylene microplastics in zebrafish: Perspective study of genotoxicity, mutagenicity, and redox unbalance.

Despite the toxicity of microplastics (MPs) in freshwater fish has been demonstrated in previous studies, their effects when mixed with other pollutants (organic and inorganic) are poorly understood. Thus, we aimed to test the hypothesis that the association of polyethylene MPs (PE-MPs) to a mix of emerging pollutants induces more adverse genotoxic, mutagenic, and redox unbalance effects in adult zebrafish (Danio rerio), after 15 days of exposure. Although the accumulation of MPs in animals was greater in animals exposed to PE-MPs alone, erythrocyte DNA damage (comet assay) and the frequency of erythrocytic nuclear abnormalities (ENAs) evidenced in zebrafish exposed to PE-MPs alone were as pronounced as those observed in animals exposed to the mix of pollutant (alone or in combination with MPs), which constitutes the big picture of the current study. Moreover, we noticed that such effects were associated with an imbalance between pro-and antioxidant metabolism in animals, whose activity of superoxide dismutase (SOD) and catalase (CAT) was assessed in different organs which were not sufficient to counterbalance the production of reactive oxygen species [hydrogen peroxide (H2O2)] and nitrogen [nitric oxide (NO)] evaluated. The principal component analysis (PCA) also revealed that while the antioxidant activity was more pronounced in the brain and liver of animals, the highest production of H2O2 was perceived in the gills and muscles, suggesting that the biochemical response of the animals was organ-dependent. Thus, the present study did not demonstrate antagonistic, synergistic, or additive effects on animals exposed to the combination between PE-MPs and a mix of pollutants in the zebrafish, which reinforces the theory that interactions between pollutants in aquatic ecosystems may be as complex as their effects on freshwater ichthyofauna.