Hidden ecotoxicological dangers: Investigating pathogen circulation and non-toxic risks hazards in a crucial brazilian watershed.

Numerous studies evaluate chemical contaminants released by human activities and their effects on biota and aquatic ecosystems. However, few of these studies address non-toxic agents and their potentially harmful effects, which, in a concealed manner, culminate in an increased ecotoxicological risk for aquatic life and public health. This study investigated the presence of toxic and non-toxic pollutants in one of the main watersheds in Northeast Brazil (Rio São Francisco) and proposed a model of dispersion and transfer of resistance among the analyzed bacteria, also assessing the health risks of individuals and aquatic organisms present there. The results are worrying because although most toxic parameters, including physical-chemical and chromatographic aspects, comply with Brazilian environmental standards, non-toxic (microbiological) parameters do not. This research reveals the circulation of pathogens in several points of this hydrographic basin, highlighting the hidden ecotoxicological potential of an aquatic environment considered unaffected by the usual patterns of toxic parameters.

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.

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.

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 novel insight on SARS-CoV-2 S-derived fragments in the control of the host immunity.

Despite all efforts to combat the pandemic of COVID-19, we are still living with high numbers of infected persons, an overburdened health care system, and the lack of an effective and definitive treatment. Understanding the pathophysiology of the disease is crucial for the development of new technologies and therapies for the best clinical management of patients. Since the manipulation of the whole virus requires a structure with an adequate level of biosafety, the development of alternative technologies, such as the synthesis of peptides from viral proteins, is a possible solution to circumvent this problem. In addition, the use and validation of animal models is of extreme importance to screen new drugs and to compress the organism's response to the disease. Peptides derived from recombinant S protein from SARS-CoV-2 were synthesized and validated by in silico, in vitro and in vivo methodologies. Macrophages and neutrophils were challenged with the peptides and the production of inflammatory mediators and activation profile were evaluated. These peptides were also inoculated into the swim bladder of transgenic zebrafish larvae at 6 days post fertilization (dpf) to mimic the inflammatory process triggered by the virus, which was evaluated by confocal microscopy. In addition, toxicity and oxidative stress assays were also developed. In silico and molecular dynamics assays revealed that the peptides bind to the ACE2 receptor stably and interact with receptors and adhesion molecules, such as MHC and TCR, from humans and zebrafish. Macrophages stimulated with one of the peptides showed increased production of NO, TNF-α and CXCL2. Inoculation of the peptides in zebrafish larvae triggered an inflammatory process marked by macrophage recruitment and increased mortality, as well as histopathological changes, similarly to what is observed in individuals with COVID-19. The use of peptides is a valuable alternative for the study of host immune response in the context of COVID-19. The use of zebrafish as an animal model also proved to be appropriate and effective in evaluating the inflammatory process, comparable to humans.

Photobiomodulation Reduces the Cytokine Storm Syndrome Associated with COVID-19 in the Zebrafish Model.

Although the exact mechanism of the pathogenesis of coronavirus SARS-CoV-2 (COVID-19) is not fully understood, oxidative stress and the release of pro-inflammatory cytokines have been highlighted as playing a vital role in the pathogenesis of the disease. In this sense, alternative treatments are needed to reduce the level of inflammation caused by COVID-19. Therefore, this study aimed to investigate the potential effect of red photobiomodulation (PBM) as an attractive therapy to downregulate the cytokine storm caused by COVID-19 in a zebrafish model. RT-qPCR analyses and protein-protein interaction prediction among SARS-CoV-2 and Danio rerio proteins showed that recombinant Spike protein (rSpike) was responsible for generating systemic inflammatory processes with significantly increased levels of pro-inflammatory (il1b, il6, tnfa, and nfkbiab), oxidative stress (romo1) and energy metabolism (slc2a1a and coa1) mRNA markers, with a pattern similar to those observed in COVID-19 cases in humans. On the other hand, PBM treatment was able to decrease the mRNA levels of these pro-inflammatory and oxidative stress markers compared with rSpike in various tissues, promoting an anti-inflammatory response. Conversely, PBM promotes cellular and tissue repair of injured tissues and significantly increases the survival rate of rSpike-inoculated individuals. Additionally, metabolomics analysis showed that the most-impacted metabolic pathways between PBM and the rSpike treated groups were related to steroid metabolism, immune system, and lipid metabolism. Together, our findings suggest that the inflammatory process is an incisive feature of COVID-19 and red PBM can be used as a novel therapeutic agent for COVID-19 by regulating the inflammatory response. Nevertheless, the need for more clinical trials remains, and there is a significant gap to overcome before clinical trials can commence.

Exposure of adult zebrafish (Danio rerio) to SARS-CoV-2 at predicted environmentally relevant concentrations: Outspreading warns about ecotoxicological risks to freshwater fish.

While the multifaceted social, economic, and public health impacts associated with the COVID-19 pandemic are known, little is known about its effects on non-target aquatic ecosystems and organisms. Thus, we aimed to evaluate the potential ecotoxicity of SARS-CoV-2 lysate protein (SARS.CoV2/SP02.2020.HIAE.Br) in adult zebrafish (Danio rerio) at predicted environmentally relevant concentrations (0.742 and 2.226 pg/L), by 30 days. Although our data did not show locomotor alterations or anxiety-like or/and anxiolytic-like behavior, we noticed that exposure to SARS-CoV-2 negatively affected habituation memory and social aggregation of animals in response to a potential aquatic predator (Geophagus brasiliensis). An increased frequency of erythrocyte nuclear abnormalities was also observed in animals exposed to SARS-CoV-2. Furthermore, our data suggest that such changes were associated with a redox imbalance [↑ROS (reactive oxygen species), ↑H2O2 (hydrogen peroxide), ↓SOD (superoxide dismutase), and ↓CAT (catalase)], cholinesterasic effect [↑AChE (acetylcholinesterase) activity], as well as the induction of an inflammatory immune response [↑NO (nitric oxide), ↑IFN-γ (interferon-gamma), and ↓IL-10 (interleukin-10)]. For some biomarkers, we noticed that the response of the animals to the treatments was not concentration-dependent. However, principal component analysis (PCA) and the "Integrated Biomarker Response" index (IBRv2) indicated a more prominent ecotoxicity of SARS-CoV-2 at 2.226 pg/L. Therefore, our study advances knowledge about the ecotoxicological potential of SARS-CoV-2 and reinforces the presumption that the COVID-19 pandemic has negative implications beyond its economic, social, and public health impacts.

Long-term exposure of zebrafish juveniles to carbon nanofibers at predicted environmentally relevant concentrations: Outspreading warns about ecotoxicological risks to freshwater fish.

Although carbon-based nanomaterials (CNMs) toxicity has already been demonstrated in some animal models, little is known about the impact of carbon nanofibers (CNFs) on aquatic vertebrates. Thus, we aimed to evaluate the possible effects of long-term exposure of zebrafish (Danio rerio) juveniles (90 days) to CNFs in predicted environmentally relevant concentrations (10 ng/L and 10 µg/L). Our data revealed that exposure to CNFs did not affect the growth and development of the animals, in addition to not having induced locomotor alterations or anxiety-like behavior. On the other hand, we observed that zebrafish exposed to CNFs showed a response deficit to the vibratory stimulus test, alteration in the density of neuromasts recorded in the final ventral region, as well as an increase in thiobarbituric acid reactive substances levels and a reduction in total antioxidant activity, nitric oxide, and acetylcholinesterase activity in the brain. Such data were directly associated with a higher concentration of total organic carbon in the brain, which suggests the bioaccumulation of CNFs. Furthermore, exposure to CNFs induced a picture suggestive of genomic instability, inferred by the increased frequency of nuclear abnormalities and DNA damage in circulating erythrocytes. Although the individual analyses of the biomarkers did not point to a concentration-dependent effect, the principal component analysis (PCA) and the Integrated Biomarker Response Index (IBRv2) indicate a more prominent effect induced by the higher CNFs concentration (10 µg/L). Therefore, our study confirms the impact of CNFs in the studied model (D. rerio) and sheds light on the ecotoxicological risks of these nanomaterials to freshwater fish. Based on the ecotoxicological screening provided by our study, new horizons are opened for investigations into the mechanisms of action of CNFs, which will help understand the magnitude of the impact of these materials on aquatic biota.

Multiple endpoints of polyethylene microplastics toxicity in vascular plants of freshwater ecosystems: A study involving Salvinia auriculata (Salviniaceae).

More recently, the number of studies on the impacts of microplastics (MPs) on plants has drawn attention considerably. However, many of these studies focused on terrestrial plants, with vascular plants from freshwater ecosystems being little studied. Thus, we aimed to evaluate the possible effects of exposure of Salvinia auriculata, for 28 days, to different concentrations of polyethylene MPs (PE MPs - diameter: 35.46 ± 18.17 µm) (2.7 ×108 and 8.1 ×108 particles/m3), using different biomarkers. Our data indicated that exposure to PE MPs caused alterations in plant growth/development (inferred by the lower floating frond number, "root" length, and the number of "roots"), as well as lower dispersion of individuals in the experimental units. Plants exposed to PE MPs also showed lower epidermal thickness (abaxial leaf face) and a longer length of the central leaf vein and vascular bundle area. Ultrastructural analyses of S. auriculata exposed to MPs revealed rupture of some epidermal cells and trichomes on the adaxial and abaxial, leaf necrosis, and chlorosis. In the "roots", we observed dehydrated filamentous structures with evident deformations in plants exposed to the pollutants. Both on the abaxial leaf face and on the "roots", the adherence of PE MPs was observed. Furthermore, exposure to PE MPs induced lower chlorophyll content, cell membrane damage, and redox imbalance, marked by reduced catalase and superoxide dismutase activity and increased production of reactive oxygen and nitrogen species as well as malondialdehyde. However, in general, we did not observe the dose-response effect for the evaluated biomarkers. The values of the integrated biomarker response index, the principal component analysis (PCA) results and the hierarchical clustering analysis confirmed the similarity between the responses of plants exposed to different PE MPs concentrations. Therefore, our study sheds light on how PE MPs can affect S. auriculata and reinforces that putting these pollutants in freshwater environments might be hazardous from an ecotoxicological point of view.

Exposure to polystyrene nanoplastics induces an anxiolytic-like effect, changes in antipredator defensive response, and DNA damage in Swiss mice.

Although the in vivo toxicity of nanoplastics (NPs) has already been reported in different model systems, their effects on mammalian behavior are poorly understood. Thus, we aimed to evaluate whether exposure to polystyrene (PS) NPs (diameter: 23.03 ± 0.266 nm) alters the behavior (locomotor, anxiety-like and antipredator) of male Swiss mice, induces brain antioxidant activity, and erythrocyte DNA damage. For this, the animals were exposed to NPs for 20 days at different doses (6.5 ng/kg and 6500 ng/kg). Initially, we did not observe any effect of pollutants on the locomotor activity of the animals (inferred via open field test and Basso mouse scale for locomotion). However, we noticed an anxiolytic-like behavior (in the open field test) and alterations in the antipredatory defensive response of mice exposed to PS NPs, when confronted with their predator potential (snake, Pantherophis guttatus). Furthermore, such changes were associated with suppressing brain antioxidant activity, inferred by lower DPPH radical scavenging activity, reduced total glutathione content, as well as the translocation and accumulation of NPs in the brain of the animals. In addition, we noted that the treatments induced DNA damage, evaluated via a single-cell gel electrophoresis assay (comet assay) applied to circulating erythrocytes of the animals. However, we did not observe a dose-response effect for all biomarkers evaluated and the estimated accumulation of PS NPs in the brain. The values of the integrated biomarker response index and the results of the principal component analysis (PCA) and the hierarchical clustering analysis confirmed the similarity between the responses of animals exposed to different doses of PS NPs. Therefore, our study sheds light on how PS NPs can impact mammals and reinforce the ecotoxicological risk associated with the dispersion of these pollutants in natural environments and their uptake by mammals.

Toxicity assessment of polyethylene microplastics in combination with a mix of emerging pollutants on Physalaemus cuvieri tadpoles.

Studies in recent years have shown that aquatic pollution by microplastics (MPs) can be considered to pose additional stress to amphibian populations. However, our knowledge of how MPs affect amphibians is very rudimentary, and even more limited is our understanding of their effects in combination with other emerging pollutants. Thus, we aimed to evaluate the possible toxicity of polyethylene MPs (PE-MPs) (alone or in combination with a mix of pollutants) on the health of Physalaemus cuvieri tadpoles. After 30 days of exposure, multiple biomarkers were measured, including morphological, biometric, and developmental indices, behavioral parameters, mutagenicity, cytotoxicity, antioxidant and cholinesterase responses, as well as the uptake and accumulation of PE-MPs in animals. Based on the results, there was no significant change in any of the parameters measured in tadpoles exposed to treatments, but induced stress was observed in tadpoles exposed to PE-MPs combined with the mixture of pollutants, reflecting significant changes in physiological and biochemical responses. Through principal component analysis (PCA) and integrated biomarker response (IBR) assessment, effects induced by pollutants in each test group were distinguished, confirming that the exposure of P. cuvieri tadpoles to the PE-MPs in combination with a mix of emerging pollutants induces an enhanced stress response, although the uptake and accumulation of PE-MPs in these animals was reduced. Thus, our study provides new insight into the danger to amphibians of MPs coexisting with other pollutants in aquatic environments.

Toxicity assessment of SARS-CoV-2-derived peptides in combination with a mix of pollutants on zebrafish adults: A perspective study of behavioral, biometric, mutagenic, and biochemical toxicity.

The dispersion of SARS-CoV-2 in aquatic environments via the discharge of domestic and hospital sewage has been confirmed in different locations. Thus, we aimed to evaluate the possible impacts of zebrafish (Danio rerio) exposure to SARS-CoV-2 peptide fragments (PSPD-2001, 2002, and 2003) alone and combined with a mix of emerging pollutants. Our data did not reveal the induction of behavioral, biometric, or mutagenic changes. But we noticed an organ-dependent biochemical response. While nitric oxide and malondialdehyde production in the brain, gills, and muscle did not differ between groups, superoxide dismutase activity was reduced in the "PSPD", "Mix", and "Mix+PSPD" groups. An increase in catalase activity and a reduction in DPPH radical scavenging activity were observed in the brains of animals exposed to the treatments. However, the "Mix+PSPD" group had a higher IBRv2 value, with NO levels (brain), the reduction of acetylcholinesterase activity (muscles), and the DPPH radical scavenging activity (brain and muscles), the most discriminant factors for this group. The principal component analysis (PCA) and hierarchical clustering analysis indicated a clear separation of the "Mix+PSPD" group from the others. Thus, we conclude that exposure to viral fragments, associated with the mix of pollutants, induced more significant toxicity in zebrafish adults than in others.

Short-term exposure of the mayfly larvae (Cloeon dipterum, Ephemeroptera: Baetidae) to SARS-CoV-2-derived peptides and other emerging pollutants: A new threat for the aquatic environments.

The input of SARS-CoV-2 or its fragments into freshwater ecosystems (via domestic or hospital sewage) has raised concerns about its possible impacts on aquatic organisms. Thus, using mayfly larvae [Cloeon dipterum (L.), Ephemeroptera: Baetidae] as a model system, we aimed to evaluate the possible effects of the combined short exposure of SARS-CoV-2-derived peptides (named PSPD-2001, PSPD-2002, and PSPD-2003 - at 266.2 ng/L) with multiple emerging pollutants at ambient concentrations. After six days of exposure, we observed higher mortality of larvae exposed to SARS-CoV-2-derived peptides (alone or in combination with the pollutant mix) and a lower-body condition index than those unexposed larvae. In the "PSPD" and "Mix+PSPD" groups, the activity of superoxide dismutase, catalase, DPPH radical scavenging activity, and the total thiol levels were also lower than in the "control" group. In addition, we evidenced the induction of nitrosative stress (inferred by increased nitrite production) and reduced acetylcholinesterase activity by SARS-CoV-2-derived peptides. On the other hand, malondialdehyde levels in larvae exposed to treatments were significantly lower than in unexposed larvae. The values of the integrated biomarker response index and the principal component analysis (PCA) results confirmed the similarity between the responses of animals exposed to SARS-CoV-2-derived peptides (alone and in combination with the pollutant mix). Although viral peptides did not intensify the effects of the pollutant mix, our study sheds light on the potential ecotoxicological risk associated with the spread of the new coronavirus in aquatic environments. Therefore, we recommend exploring this topic in other organisms and experimental contexts.

Shedding light on the toxicity of SARS-CoV-2-derived peptide in non-target COVID-19 organisms: A study involving inbred and outbred mice.

Despite advances in research on the vaccine and therapeutic strategies of COVID-19, little attention has been paid to the possible (eco)toxicological impacts of the dispersion of SARS-CoV-2 particles in natural environments. Thus, in this study, we aimed to evaluate the behavioral and biochemical consequences of the short exposure of outbred and inbred mice (male Swiss and C57Bl/6 J mice, respectively) to PSPD-2002 (peptide fragments of the Spike protein of SARS-CoV-2) synthesized in the laboratory. Our data demonstrated that after 24 h of intraperitoneal administration of PSPD-2002 (at 580 µg/kg) the animals did not present alterations in their locomotor, anxiolytic-like, or anxiety-like behavior (in the open field test), nor antidepressant-like or depressive behavior in the forced swimming test. However, the C57Bl/6 J mice exposed to PSPD-2002 showed memory deficit in the novel object recognition task, which was associated with higher production of thiobarbituric acid reactive substances, as well as the increased suppression of acetylcholinesterase brain activity, compared to Swiss mice also exposed to peptide fragments. In Swiss mice the reduction in the activity of superoxide dismutase and catalase in the brain was not associated with increased oxidative stress biomarkers (hydrogen peroxide), suggesting that other antioxidant mechanisms may have been activated by exposure to PSPD-2002 to maintain the animals' brain redox homeostasis. Finally, the results of all biomarkers evaluated were applied into the "Integrated Biomarker Response Index" (IBRv2) and the principal component analysis (PCA), and greater sensitivity of C57Bl/6 J mice to PSPD-2002 was revealed. Therefore, our study provides pioneering evidence of mammalian exposure-induced toxicity (non-target SARS-CoV-2 infection) to PSPD-2002, as well as "sheds light" on the influence of genetic profile on susceptibility/resistance to the effects of viral peptide fragments.

Gdnf Acts as a Germ Cell-Derived Growth Factor and Regulates the Zebrafish Germ Stem Cell Niche in Autocrine- and Paracrine-Dependent Manners.

Glial cell line-derived neurotrophic factor (GDNF) and its receptor (GDNF Family Receptor α1-GFRα1) are well known to mediate spermatogonial stem cell (SSC) proliferation and survival in mammalian testes. In nonmammalian species, Gdnf and Gfrα1 orthologs have been found but their functions remain poorly investigated in the testes. Considering this background, this study aimed to understand the roles of the Gdnf-Gfrα1 signaling pathway in zebrafish testes by combining in vivo, in silico and ex vivo approaches. Our analysis showed that zebrafish exhibit two paralogs for Gndf (gdnfa and gdnfb) and its receptor, Gfrα1 (gfrα1a and gfrα1b), in accordance with a teleost-specific third round of whole genome duplication. Expression analysis further revealed that both ligands and receptors were expressed in zebrafish adult testes. Subsequently, we demonstrated that gdnfa is expressed in the germ cells, while Gfrα1a/Gfrα1b was detected in early spermatogonia (mainly in types Aund and Adiff) and Sertoli cells. Functional ex vivo analysis showed that Gdnf promoted the creation of new available niches by stimulating the proliferation of both type Aund spermatogonia and their surrounding Sertoli cells but without changing pou5f3 mRNA levels. Strikingly, Gdnf also inhibited late spermatogonial differentiation, as shown by the decrease in type B spermatogonia and down-regulation of dazl in a co-treatment with Fsh. Altogether, our data revealed that a germ cell-derived factor is involved in maintaining germ cell stemness through the creation of new available niches, supporting the development of spermatogonial cysts and inhibiting late spermatogonial differentiation in autocrine- and paracrine-dependent manners.

Toxicity induced via ingestion of naturally-aged polystyrene microplastics by a small-sized terrestrial bird and its potential role as vectors for the dispersion of these pollutants.

In recent years, there has been a growing number of studies on the impact of microplastics (MPs) on biota. However, its effects on birds' health are poorly understood. Thus, we aimed to evaluate the possible effects of ingestion of naturally-aged MPs by Coturnix Coturnix japonica (11 and 22 MP particles/day/bird, once a day, for 9 days), from different toxicity biomarkers. At the end of the experiment, it was found that the ingested MPs in birds showed a significant reduction in body biomass. Also, an increase in malondialdehyde production in the liver, brain, intestine, and gizzard of the birds, as well as a suppressive effect on hepatic nitric oxide production and superoxide dismutase activity in the liver and intestine were observed. Cerebral catalase activity was reduced in birds exposed to MPs and the cholinesterasic effect (marked by increased acetylcholinesterase activity) was observed in the muscle and brain of these animals. Despite these differences, through the main component analysis, hierarchical clustering analysis, and integrated biomarker response assessment, we observed similar toxicological effects in birds exposed to different amounts of MPs. In addition, the size of MPs was reduced, and their shape was altered as they transited through the gastrointestinal system, which probably explains their accumulation in the liver of birds. An expressive number of MPs are released through the feces of the birds throughout the experiment. As far as we know, this is the first report that associates MPs ingestion by small-sized terrestrial birds with biochemical alterations viz., predictive of oxidative stress, redox imbalance, and cholinesterasic effect, in addition to shedding light on the potential role of these birds as vectors for dispersal of MPs in natural environments.

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.