Integration of a battery of biomarkers to evaluate the health status of field-collected frogs of Leptodactylus luctator living in ecosystems with different anthropogenic disturbances.

Amphibians are the most threatened group of vertebrates because they have certain biological and ecological characteristics that make them sensitive to environmental changes. The aim of this study was to evaluate the health status of field-collected adult frogs of Leptodactylus luctator (Amphibia, Anura) living in sites with different anthropogenic disturbances (florihorticulture, petrochemical industry and sewage discharges) and a reference site without any detectable influence of such activities. To this end, a battery of 21 biomarkers (hematological, biochemical and individual biomarkers) was studied using a multivariate approach that allows us to evaluate the relationship between them and provide information on their usefulness. The frogs at the florihorticulture, petrochemical and sewage discharges sites exhibited several biomarkers far from homeostasis. In addition, we identified 11 of 21 biomarkers that were useful indicators of the health status of the frogs and allowed discrimination between study sites in the following order: lymphocytes (98 %), neutrophils (45 %), hemoglobin (42 %), monocytes (41 %), fat body index (35 %), eosinophils (35 %), hepatosomatic index (33 %), mean corpuscular hemoglobin (32 %), thrombocytes (27 %), catalase in liver (26 %), and GST in liver (26 %). The results suggest that hematological biomarkers contribute the most to site separation, whereas biochemical biomarkers contribute the least. The integral interpretation of the results also allowed us to diagnose the different health status of L. luctator: The frogs from the petrochemical industry were the most negatively affected, followed by the frogs from the sewages discharges and finally the frogs from the florihorticulture and reference sites. This is the first field study with anurans in which so many biomarkers were examined.

First evidence of environmental bioaccumulation of pharmaceuticals on adult native anurans (Rhinella arenarum) from Argentina.

The presence of pharmaceutically active compounds (PhACs) in surface water is well known, whereas their natural occurrence in biota is much less explored. The aim of this work was to evaluate the bioaccumulation of PhACs in adult toads of the neotropical species Rhinella arenarum. Three sites were selected in Buenos Aires (Argentina): a reference site (Site 1), a site with direct discharge from a secondary wastewater treatment plant (WWTP) (Site 2) and a site 300 m downstream of the WWTP discharge (Site 3). Surface water samples, as well as muscle, liver and fat bodies of toads were collected, extracted and analyzed by LC-MS/MS. Highly significant differences in total PhACs concentration in surface water (p < 0.005) were detected between Site 2 and the other sites. These concentrations ranged from 0.37 to 52.46 ng/L at Site 1, 0.71-6950.37 ng/L at Site 2, and 0.12-75.45 ng/L at Site 3. In general, bioaccumulation of PhACs in toad tissues was similar between sites and tissues of each site. The highest concentrations were detected in the muscle of toads from Site 3 (1.06-87.24 ng/g dw), followed by liver (1.77-38.10 ng/g dw) and fat bodies (0.68-20.59 ng/g dw) from Site 1. Ibuprofen (6950 ng/L), acetaminophen (3277 ng/L) and valsartan (2504 ng/L) were the compounds with the highest concentrations in surface water from Site 2, whereas acetaminophen (87.2 ng/g dw, muscle from Site 3), desloratadine (38.1 ng/g dw, liver from Site 1), and phenazone (25.9 ng/g dw, liver from Site 1) were the ones that showed the highest concentrations in biota. This is the first time a field study has examined the environmental bioaccumulation of PhACs in anurans, demonstrating their potential for monitoring the status of natural ecosystems.

Tadpoles of the horned frog Ceratophrys ornata exhibit high sensitivity to chlorpyrifos for conventional ecotoxicological and novel bioacoustic variables.

Previous studies reported that some species of the family Ceratophryidae are able to produce sounds during premetamorphic tadpole stages. We have now determined the effects of the cholinesterase-inhibiting insecticide chlorpyrifos (CPF) on sounds emitted by tadpoles of Ceratophrys ornata. Tadpoles were exposed individually in order to evaluate the progression of effects. Effects on sound production were complemented with common ecotoxicological endpoints (mortality, behavior, abnormalities and growth inhibition). C. ornata was found to be more sensitive than other native (= 67%, 50%) and non-native species (= 75%, 100%) considering lethal and sublethal endpoints, respectively. Effects on sounds appear along with alterations in swimming, followed by the presence of mild, then severe abnormalities and finally death. Therefore, sound production may be a good biomarker since it anticipates other endpoints that are also affected by CPF. Ceratophrys ornata is a promising new model species in ecotoxicology.