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.
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•Combined embryotoxicity of nanoparticles and microplastics on bullfrog embryos.•Successful application of the FETAX protocol for bullfrog embryos.•Polyethylene microplastics caused abdominal edemas in bullfrog embryos.•TiO2 nanoparticles reduced embryos' survival and hatching rates.•Combined exposure reduced the embryotoxicity on bullfrog embryos.
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.
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•State-of-the-art review on the ecotoxicity of plastic microfibers (PMF) on aquatic organisms.•PMFs as an emerging risk factor for aquatic organisms.•Fish, Arthropoda and mussels are overrepresented in studies of PMF ecotoxicity.•Environmental concerns about PMF produced during the pandemic period.
Iron oxide nanoparticles (IONPs) are used in several medical and environmental applications, but their mechanism of action and hazardous effects to early developmental stages of fish remain unknown. ...Thus, the present study aimed to assess the developmental toxicity of citrate-functionalized IONPs (γ-Fe2O3 NPs), in comparison with its dissolved counterpart, in zebrafish (Danio rerio) after static and semi-static exposure. Embryos were exposed to environmental concentrations of both iron forms (0.3, 0.6, 1.25, 2.5, 5 and 10 mg L−1) during 144 h, jointly with negative control group. The interaction and distribution of both Fe forms on the external chorion and larvae surface were measured, following by multiple biomarker assessment (mortality, hatching rate, neurotoxicity, cardiotoxicity, morphological alterations and 12 morphometrics parameters). Results showed that IONPs were mainly accumulated on the zebrafish chorion, and in the digestive system and liver of the larvae. Although the IONPs induced low embryotoxicity compared to iron ions in both exposure conditions, these nanomaterials induced sublethal effects, mainly cardiotoxic effects (reduced heartbeat, blood accumulation in the heart and pericardial edema). The semi-static exposure to both iron forms induced high embryotoxicity compared to static exposure, indicating that the nanotoxicity to early developmental stages of fish depends on the exposure system. This is the first study concerning the role of the exposure condition on the developmental toxicity of IONPs on fish species.
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•Differential developmental toxicity of γ-Fe₂O₃ NPs and iron ions in zebrafish.•Embryotoxicity of γ-Fe₂O₃ NPs depends on the exposure condition (static and semi-static).•Semi-static exposure induced high embryotoxicity than static exposure.•γ-Fe₂O₃ NPs induced several cardiotoxic effects on zebrafish embryos.
The environmental health impact of titanium dioxide nanoparticles (TiO
2
NPs) is of emerging concern, but their effects and ecotoxicity resulting from the interaction between TiO
2
NPs and metals, ...such as cadmium (Cd), on aquatic organisms remain limited. Thus, the current study aimed to evaluate the ecotoxic effects of TiO
2
NPs and Cd (alone or in co-exposure) on zebrafish embryos and larvae, based on using multiple biomarkers. Embryos were exposed to environmentally relevant TiO
2
NPs (0.1, 1.0, and 10 µg L
−1
) and Cd (10 µg L
−1
) concentrations, either alone or in a mix, for 168 h, whereas the control group was kept in reconstituted water. Results showed that the isolated TiO
2
NPs have low toxicity to zebrafish embryos and larvae. However, Cd — alone or in co-exposure with TiO
2
NPs — induced toxic effects in zebrafish, such as cardiotoxicity, as well as morphological and morphometric changes. The present study has shown TiO
2
NPs/Cd interactive effects, including decreased toxicity, on the assessed models. Results have evidenced the potential environmental risk of the nanoparticle and metal mixtures, mainly at early vertebrate developmental stages.
Graphical abstract
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 (TiO
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 TiO
NPs, and to a lesser extent, also affected the Mix group. TiO
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.
Dentre os répteis, os lagartos e os testudines são o foco de inúmeras investigações que contemplam o estudo de sua ontogenia, contudo, descrições do desenvolvimento e da sequência de ossificação em ...crocodilianos são escassas. Assim, objetivou-se investigar o padrão de formação e a sequência de ossificação das vértebras e costelas em Caiman yacare. Embriões foram coletados em intervalos regulares e submetidos ao protocolo de diafanização e coloração por Aliarina red S. O processo de ossificação da coluna vertebral ocorre no sentido crânio-caudal a partir dos 33 dias. As vértebras se formam a partir de centros de ossificação distintos para os centros, o arco e as apófises. O pró-atlas se forma a partir de dois centros de ossificação distintos que se fundem até o fim do período de incubação. A fusão dorsal dos processos transversos ocorre parcialmente aos 51 dias, dando origem ao diminuto esboço do processo espinhoso. No embrião de 57 dias a articulação entre as costelas, o sacral e o ílio não estava evidente. A morfologia e tamanho das vértebras caudais variam consideravelmente no sentido crânio-caudal. Todas apresentam os processos hemais e iniciam sua ossificação até os 48 dias. C. yacare apresenta um número variado de costelas abdominais, que se ossificam ainda durante o período embrionário. O padrão da esqueletogênese de C. yacare é congruente com de outros répteis, embora existam algumas variações, com particular ênfase na remodelação de estruturas, o que possivelmente reflete as variações do período de incubação entre os diversos répteis relatados.
Pesticides are considered one of the main causes of the population decline of reptiles worldwide, with freshwater turtles being particularly susceptible to aquatic contamination. In this context, we ...investigated the potential mutagenic, hepatotoxic, and neurotoxic effects in neonates of Podocnemis expansa exposed to substrate contaminated with different concentrations of glyphosate and/or fipronil during embryonic development. Eggs collected from the natural environment were artificially incubated in sand moistened with pure water, water added with glyphosate Atar 48® at concentrations of 65 and 6500 μg/L (groups G1 and G2, respectively), water added with fipronil Regent® 800WG at 4 and 400 μg/L (groups F1 and F2, respectively) and, water added with the combination of 65 μg/L glyphosate and 4 μg/L fipronil or with 6500 μg/L glyphosate and 400 μg/L fipronil (groups GF1 and GF2, respectively). For mutagenicity analysis, we evaluated the frequency of micronuclei (MN) and other erythrocyte nuclear abnormalities (ENAs), while for evaluation of hepatotoxicity and neurotoxicity, livers and encephalon were analyzed for histopathological alterations. Exposure to pesticides, alone or in combination, increased the frequency of erythrocyte nuclear abnormalities, particularly blebbed nuclei, moved nuclei, and notched nuclei. Individuals exposed to fipronil exhibited congestion and inflammatory infiltrate in their liver tissue, while, in the encephalon, congestion, and necrosis were present. Our study confirms that the incubation of eggs in substrate polluted with glyphosate and fipronil causes histopathological damage and mutagenic alteration in P. expansa, highlighting the importance of using different biomarkers to evaluate the ecotoxicological effects of these pesticides, especially in oviparous animals.
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•Exposure to pesticides, alone or in combination, increased the frequency of erythrocyte nuclear abnormalities.•Individuals exposed to pesticides showed histopathological changes in the liver and brain.•Incubation of eggs in substrate polluted with glyphosate and fipronil causes damage in P. expansa.
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 (ZnCl
2
) 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 ZnCl
2
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.
Graphical abstract
Besides its crucial role during embryo development, eggshells are an essential bioindicator of environmental contaminants. However, the effects of contaminant exposure during incubation on the ...eggshell composition remain poorly known for freshwater turtles. Accordingly, we tested the effects of incubating the eggs of Podocnemis expansa in substrates containing glyphosate and fipronil formulations on the eggshell's level of mineral and dry matter, crude protein, nitrogen, and ethereal extract. Eggs were incubated in sand moistened with water contaminated with glyphosate Atar 48 at concentrations of 65 or 6500 μg/L, fipronil Regent 800 WG at concentrations of 4 or 400 μg/L, or the combination of 65 μg/L glyphosate and 4 μg/L fipronil and 6500 μg/L glyphosate with 400 μg/L fipronil. Exposure to the tested pesticides, alone or in association, altered the chemical composition of the eggshell of P. expansa, reducing the eggshell's moisture and crude protein content and increasing levels of ethereal extract. These changes may cause significant deficiencies in the mobilization of water and nutrients to the embryo, affecting the development and reproductive success of P. expansa.
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•Eggshell composition of Amazon turtle after exposure to fipronil and glyphosate is evaluated.•Exposure to the pesticides, alone or in association, altered the chemical composition of the eggshell of P. expansa.•Fipronil and glyphosate reduced the eggshell's moisture and crude protein content.•Tested pesticide-induced increase of ethereal extract levels in eggshell of Amazon turtle.
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