A realistic combined exposure scenario: effect of microplastics and atrazine on Piaractus mesopotamicus.

Microplastics, considered emerging environmental contaminants resulting from plastic degradation, are discovered in diverse aquatic ecosystems and can be unintentionally ingested by fish. Therefore, it is essential to characterize their interaction with other contaminants, such as agrochemicals, in aquatic environments. This study aimed to assess histological, enzymatic, and genotoxic biomarkers in juvenile pacú (Piaractus mesopotamicus) exposed to polyethylene (PE) microplastic particles and the herbicide atrazine, individually or combined, for 15 days. Four treatments were used: a negative control (CON), PE in the fish diet (0.1% w/w, FPE), atrazine through water (100 µg L-1, ATZ), and the mixture (ATZ+FPE). Results confirmed histological alterations in gills (edema and lamellar fusion) and liver (necrotic areas and congestion) of fish exposed to ATZ and ATZ+FPE. The number of goblet cells increased in the posterior intestine of fish under ATZ+FPE compared to CON and FPE. Enzyme activities (CAT, GST, AChE, and BChE) significantly increased in ATZ+FPE compared to CON. However, no genotoxic effect was demonstrated. These findings provide insights into the complex impacts of simultaneous exposure to atrazine and microplastics, emphasizing the need for continued research to guide effective environmental management strategies against these contaminants that represent a risk to aquatic organisms.

Anuran heart development and critical developmental periods: A comparative analysis of three neotropical anuran species.

The heart begins to form early during vertebrate development and is the first functional organ of the embryo. This study aimed to describe and compare the heart development in three Neotropical anuran species, Physalaemus albonotatus, Elachistocleis bicolor, and Scinax nasicus. Different Gosner Stages (GS) of embryos (GS 18-20) and premetamorphic (GS 21-25), prometamorphic (GS 26-41), and metamorphic (GS 42-46) tadpoles were analyzed using stereoscopic microscopy and Scanning Electronic Microscopy. Heart development was similar in the three analyzed species; however, some heterochronic events were identified between P. albonotatus and S. nasicus compared to E. bicolor. In addition, different patterns of melanophores arrangement were observed. During the embryonic and metamorphic periods, the main morphogenetic events occur: formation of the heart tube, regionalization of the heart compartments, development of spiral valve, onset of heartbeat, looping, and final displacement of the atrium and its complete septation. Both periods are critical for the normal morphogenesis and the correct functioning of the anuran heart. These results are useful to characterize the normal anuran heart morphology and to identify possible abnormalities caused by exposure to environmental contaminants.

Biotoxicity of diclofenac on two larval amphibians: Assessment of development, growth, cardiac function and rhythm, behavior and antioxidant system.

The non-steroidal anti-inflammatory drug diclofenac (DCF) threatens the health of aquatic animals and ecosystems. In the present study, different biological endpoints (mortality, development and growth, abnormalities, cardiotoxicity, neurotoxicity and antioxidant system) were used to characterize the acute and chronic effects of DCF (at concentrations ranging between 125 and 4000 µg L-1) on two amphibian species from Argentina (Trachycephalus typhonius and Physalaemus albonotatus). Results showed that the larval developmental, growth rates, and body condition of DCF-exposed individuals of both species were significantly reduced. DCF-exposed individuals also showed several morphological abnormalities, including significantly altered body axis, chondrocranium and hyobranchial skeleton, and organ and visceral abnormalities including cardiac hypoplasia, malrotated guts, asymmetrically inverted guts, and cholecystitis. DCF also had a significant effect on the swimming performance of both species: at low concentrations (125 and 250 µg L-1), swimming distance, velocity and global activity decreased, whereas, at high concentrations (1000 and 2000 µg L-1), these behavioral responses increased. Regarding cardiac function and rhythm, at DCF concentrations higher than 1000 µg L-1, the heart frequency and ventricular systole interval of both species were significantly reduced. Regarding the antioxidant system, the activity of acetylcholinesterase indicated that DCF is neurotoxic and thus related to the changes in behavioral performance. The DCF concentrations studied produced a biochemical imbalance between radical oxygen species production and antioxidant systems. The sensitivities to sublethal and chronic DCF exposure in both anuran species were similar, thus indicating the inherent complexity involved in understanding the biotoxic effects of DCF.

Histological and immunohistochemical characterization of the integument and parotoids glands Rhinella bergi (Anura: Bufsonidae): Development and differentiation.

A detailed description of the tegument and parotoid glands of pre-metamorphic, post-metamorphic, juvenile and adult individuals of Rhinella bergi is presented to provide an exhaustive analysis of the integumentary characteristics of this species. Fragments of the tegument were fixed in Bouin solution and preserved in buffered Formol 10%. Subsequently, scanning electron microscopy (SEM) was performed to characterize the macroscopic structure of these regions. Microscopic observations were made from histological sections stained with Hematoxylin and Eosin, Alcian Blue (pH 2,5), PAS-H, Coomassie Blue, Oil Red, and Bielschowsky Impregnation.. There were three types of protuberance: warts, tubers, and thorns. These structures became evident from post-metamorphic stages. The ventral surface shows elevations similar to flat warts; however, tubers and spines are absent. Histologically, each structure consists of a spongy dermis of lax connective tissue and dense and compact dermis, associated with granular glands and a keratinized epidermis. The latter, in the dorsal region, forms projections called thorns. The granular glands accumulate, and their alveoli increase in size progressively. This work provides a morphological and histological description of the integument and the parotoid glands during the larval and post-metamorphic stage of the genus Rhinella, with aspects described for the first time in the genus.

MELANOMACRÓFAGOS HEPÁTICOS Y ACTIVIDAD DE LA ENZIMA GLUTATION S-TRANSFERASA EN Leptodactylus chaquensis (ANURA, LEPTODACTYLIDAE) COMO BIOMARCADORES DE ESTRÉS OXIDATIVO POR LA EXPOSICIÓN A CLORPIRIFOS / Liver Melanomacrophages and Gluthation S-Transferase Activity in Leptodactylus chaquensis (Anura, Leptodactylidae) as Biomarkers of Oxidative Stress Due to Chlorpyrifos Exposition

RESUMEN Se cuantificaron y compararon los melanomacrófagos (MM) y la actividad de la enzima Glutatión S-Transferasa (GST) en hígados de ejemplares adultos de Leptodactylus chaquensis (Anura, Leptodactylidae) colectados en un cultivo de arroz (CA) (siete días posteriores a la aplicación de clorpirifos, CPF) y en un sitio de referencia (SR) en el departamento San Javier, provincia de Santa Fe (Argentina). El análisis histológico reveló una mayor cantidad (p = 0.028) y área ocupada por MM (p = 0.017) y se registró una inhibición en la actividad de la GST (p = 0.030) en individuos del CA respecto a los del SR. Los efectos descriptos evidencian el riesgo ecotoxicológico para los anuros en los cultivos de arroz con aplicaciones de CPF.

ABSTRACT We quantified and compared the hepatic melanomacrophage (MM) and glutathione S-transferase (GST) enzyme activity (two oxidative stress biomarkers) in the liver of Leptodatylus chaquensis adults (Anura, Leptodactylidae) collected in a rice field (CA) in San Javier department, Santa Fe (Argentina), seven days after the application of chlorpyrifos and in a reference site (SR). The histological analysis revealed a significant amount (p = 0.028) and area occupied by MM (p = 0.017) in livers of CA compared to SR. Furthermore, a significant inhibition of GST activity was recorded in the CA frogs compared to the SR (p = 0.030). The histopathological and enzymatic effects provide evidences of ecotoxicological risk for anurans in rice field with CPF application.

Ecotoxicity of veterinary enrofloxacin and ciprofloxacin antibiotics on anuran amphibian larvae.

The ecological risks posed by two ß-diketone antibiotics (DKAs, enrofloxacin, ENR and ciprofloxacin, CPX), characterized by their long persistence in aqueous environments and known deleterious effect on model organisms such as zebrafish were analysed using Rhinella arenarum larvae. Sublethal tests were conducted using environmentally relevant concentrations of both ENR and CPX (1-1000µgL-1) under standard laboratory conditions for 96h. Biological endpoints and biomarkers evaluated were body size, shape, development and growth rates, and antioxidant enzymes (glutathione-S-transferase, GST; Catalase, CAT). Risk assessment was analysed based on ration quotients (RQ). The size and shape measurements of the larvae exposed to concentrations greater than 10µgL-1 of CPX were lower compared to controls (Dunnett post hoc p<0.05) and presented signs of emaciation. Concentrations of 1000µgL-1of CPX induced GST activity, in contrast with inhibited GST and CAT of larvae exposed to ENR. Risk assessments indicated that concentrations greater than or equal to10µgL-1 of CPX and ENR are ecotoxic for development, growth, detoxifying, and oxidative stress enzymes. It is suggested that additional risk assessments may provide evidence of bioaccumulation of CPX and ENR in tissues or organs of amphibian larvae by mesocosm sediment test conditions. Finally, intestinal microbiome studies should be considered to establish the mechanisms of action of both antibiotics.