Histopathological Evaluation of Combined Impacts of the Synthetic Progestin Levonorgestrel and Temperature on the Female Zebrafish Maturation Using a Semi-quantitative Grading AnalysisIs it Enough?

Pharmaceuticals contamination (e.g., synthetic progestins), and global climate change, represent two of the most stressful factors affecting aquatic species. To our knowledge, there is huge gap of data regarding the combined effects of both stressors on vertebrates' reproduction. Thus, it is crucial to implement rapid screenings of measurable histopathological alterations in fish gonads. For that, we propose: (1) an evaluation of the combined effects of progestin (levonorgestrel) and temperature on maturation of zebrafish female gonads, using a semi-quantitative method (i.e., grading) and (2) testing the robustness of the grading analysis comparatively to a quantitative method (i.e., stereology). Grading analysis showed a decrease on maturation stage of ovaries exposed to both stressors. Although grading is less robust than stereological analysis, it is recommended for a preliminary approach, since it gives a correct idea on trends and it is fast and cost-effective. For a detailed histological assessment, we recommend a stereological study.