Fish Embryo Toxicity Test (FET) to evaluate atrazine effects

Abstract

Surface water samples from São Paulo state were collected to perform Bioluminescent Yeast Estrogen Screen (BLYES) and chemical analyses (LC-MS/MS). Results showed environmental concentrations of atrazine from 2 to 43 ng L-1 on chemical analyses. Some studies have been performed to evaluate toxic effects on non-target organisms (fish) using herbicides such as Atrazine, a moderately toxic compound classified as an endocrine disrupting chemical (EDC) that can affect reproduction of several aquatic organisms with a compromise of vitellogenin production. To determine toxicity on embryonic stages of fish to different environmental chemicals and waste water, Fish Embryo Toxicity Test (FET) was designed using Danio rerio as model specie on this test, according to OECD 236 or ISO 15088 protocols, however these protocols observe only acute toxicity based on endpoints such as coagulated eggs, nondetachment of the tail, lack of heart beating and lack of somite formation. Some abnormalities can be recorded after the exposure on FET test but they are not considered as endpoint, neither any other compromised biomarker by EDC action. In order to evaluate the possibility of using these chronic endpoints and to verify if those environmental concentrations of atrazine are ecologically relevant, compromising reproductive aspects, FET test using Danio rerio were performed to assess lethal concentrations, sublethal concentrations and vitellogenin quantification after atrazine exposure. Occurrence of morphological abnormalities (microcephaly, spine curvature, edema, reduced size) and mortality of the embryos were determined exposing 20 fertilized eggs to atrazine concentrations from 2 to 64 mg L-1. The LC50 and EC50 were obtained after 96 hours of exposure. Organisms that survived each concentration were frozen to further vitellogenin quantification. Preliminary average concentrations obtained (LC 50; 96h= 48.15 mg L-1 and EC 50; 96h= 27 mg L-1) were considerably higher than concentrations observed on environmental samples. Therefore, surface water concentrations would not cause mortality or deformity in fish emphasizing the necessity to observe possible effect on vitellogenin concentration. Data will be analyzed and compared with the environmental concentration of atrazine to stablish the potential application of vitellogenin as endpoint on FET test.