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.

Embryonic development of Columba livia(Aves: Columbiformes) from an altricial-precocial perspective / Desarrollo embrionario de Columba livia(Aves: Columbiformes) desde una perspectiva altricial-precoz / Desenvolvimento embrionário de Columba livia(Aves: Columbiformes) a partir de uma perspectiva altricial-precoce

Objective: this study characterized the morphogenesis of Columba livia and proposed a table with 43 embryonic stages. The ontogeny of this species was divided into three phases: early, middle and late. Methods and Results: the early phase (stages 1-26) includes the final period of segmentation, gastrulation, neurulation and somitogenesis, organization of the extraembryonic membranes, and the initial formation of the major organic systems. The middle phase (stages 27-36) was characterized mainly by the growth of limbs and the organization of the autopodium, the formation and growth of the peak, and the development of integumentary annexes such as feather germs and leg scales. The late phase (stages 37-43) shows the overall embryo growth, the final organization of the pterilosis pattern and growth of feather germs, as well as final consumption of the yolk, and hatching. Conclusions: these results allow making comparisons with the proposed developmental events of Gallus gallus domesticus, highlighting similarities and differences in the ontogenetic sequence of both species. Therefore we propose some hypotheses about possible heterochronic events related to altricial-precocial developmental models.

Objetivo: en el presente trabajo se caracteriza la morfogénesis de Columba livia y se propone una tabla con 43 estadios embrionarios. La ontogenia de dicha especie fue dividida en tres etapas: temprana, media y tardía. Métodos y Resultados: la etapa temprana (estadios 1-26) incluye el periodo final de la segmentación, la gastrulación, neurulación y somitogénesis, la organización de las membranas extraembrionarias y la formación de los esbozos de los principales sistemas de órganos. La etapa media (estadios 27-36) se caracteriza principalmente por el crecimiento de los miembros y la organización del autopodio, la formación y crecimiento del pico y el desarrollo de los anexos tegumentarios como plumones y escamas de las patas. En la etapa tardía (estadios 37-43) se evidencia el crecimiento general del embrión, la organización final del patrón de pterilosis y crecimiento de plumones, como así también el consumo final del vitelo y la eclosión. Conclusiones: en base a los resultados obtenidos se realizan algunas comparaciones con respecto a los eventos del desarrollo propuestos para Gallus gallus domesticus, destacando semejanzas y diferencias en la secuencia ontogénetica de ambas especies. A partir de esto se plantean algunas hipótesis acerca de posibles eventos heterocrónicos relacionados con los modelos de desarrollo altricial-precoz.

Objetivo: no presente trabalho caracteriza-se a morfogênese de Columba livia, mediante uma tabela com 43 estágios embrionários. A ontogenia desta espécie foi dividida em três etapas: inicial, intermediária e final. Métodos y Resultados: a etapa inicial (estágios 1-26) incluiu o período final de segmentação, gastrulação, neurulação e somitogênese, organização das membranas extraembrionárias e formação dos esboços dos principais sistemas de órgãos. A etapa intermediária (estágios 27-36) foi caracterizada principalmente pelo crescimento dos membros e organização do autopódio, formação e crescimento do bico e desenvolvimento de anexos tegumentários como as penas e as escamas das patas. Na etapa tardia (estágios 37-43) foi caracterizada pelo crescimento geral do embrião, a organização final do padrão de pterilose e o padrão de crescimento das penas, assim como o consumo final do vitelo e eclosão. Conclusões: com base nos resultados obtidos, foram realizadas algumas comparações com respeito aos eventos do desenvolvimento proposto para Gallus gallus domesticus, destacando semelhanças e diferenças durante a sequência ontogenética de ambas espécies. A partir disso, algumas hipóteses são propostas acerca dos possíveis eventos heterocrônicos relacionados com os modelos de desenvolvimento altricial e precoce.

Stage-dependent toxicity of 2,4-dichlorophenoxyacetic on the embryonic development of a South American toad, Rhinella arenarum.

The acute and short term chronic toxicity of both the herbicide butyl ester of 2,4-Dichlorophenoxyacetic acid (2,4-D) and a commercial formulation (CF) were evaluated on Rhinella (= Bufo) arenarum embryos at different developmental stages. Adverse effects were analyzed by means of the isotoxicity curves for lethality, malformations, stage-dependent susceptibility, and ultrastructural features. For all experimental conditions, the CF was more toxic, up to 10 times, than the active ingredient, being the open mouth stage (S.21) the most susceptible to the herbicide. For continuous treatment conditions, the early embryonic development was the most susceptible to 2,4-D and the LC50s for 96 and 168 h were 9.06 and 7.76 mg L(-1) respectively. In addition, both the active ingredient and the CF were highly teratogenic, resulting in reduced body size, delayed development, microcephaly, agenesis of gills, and abnormal cellular proliferation processes as the main adverse effects. According to US EPA, 2,4-D in agricultural scenarios may be up to three times higher than the NOEC values for teratogenic effects reported in this study. Therefore, they might represent a risk for amphibians. This study also points out the relevance of reporting the susceptibility of embryos at different developmental stages to both the active ingredient and the CF of agrochemicals in order to protect nontarget organisms.

Stage-dependent teratogenic and lethal effects exerted by ultraviolet B radiation on Rhinella (Bufo) arenarum embryos.

The adverse effects of ultraviolet B radiation from 547.2 to 30,096 J/m2 on morphogenesis, cell differentiation, and lethality of amphibian embryos at six developmental stages were evaluated from 24 up to 168 h postexposure. The ultraviolet B radiation lethal dose 10, 50, and 90 values were obtained for all developmental stages evaluated. The lethal dose 50 values, considered as the dose causing lethality in the 50% of the organisms exposed, in J/m2 at 168 h postexposure, ranged from 2,307 to 18,930; gill circulation and blastula were the most susceptible and resistant stages, respectively. Ultraviolet B radiation caused malformations in all developmental stages but was significantly more teratogenic at the gill circulation and complete operculum stages. Moreover, at the gill circulation stage, even the lowest dose (547.2 J/m2) resulted in malformations to 100% of embryos. The most common malformations were persistent yolk plug, bifid spine, reduced body size, delayed development, asymmetry, microcephaly and anencephaly, tail and body flexures toward the irradiated side, agenesia or partial gill development, abnormal pigment distribution, and hypermotility. The stage-dependent susceptibility to ultraviolet B radiation during amphibian embryogenesis could be explained in the framework of evoecotoxicology, considering ontogenic features as biomarkers of environmental signatures of living forms ancestors during the evolutionary process. The stage-dependent susceptibility to ultraviolet B radiation on Rhinella (Bufo) arenarum embryos for both lethal and teratogenic effects could contribute to a better understanding of the role of the increased ultraviolet B radiation on worldwide amphibian populations decline.