Imazalil and prochloraz toxicokinetics in fish probed by a physiologically based kinetic (PBK) model.

Azole fungicides are highly suspected endocrine disruptors (EDs) and are frequently detected in surface water. Among them, there are prochloraz (PCZ), a commonly used  molecule for ED studies, and imazalil (IMZ), a highly suspected ED. Little is known about their toxicokinetic (TK) behavior in fish. Hence, research suggested that an improved risk assessment could be achieved by gaining insight into their TK behavior. The aim of this study is to understand and model the TK of both substances in different fish species, irrespective of the scheme of exposure. TK data from the literature were retrieved including different modes of exposure (per os and waterborne). In addition, two experiments on zebrafish exposed to either IMZ or PCZ were performed to address the lack of in vivo TK data. A physiologically based kinetic (PBK) model applied to IMZ and PCZ was developed, capable of modeling different exposure scenarios. The parameters of the PBK model were simultaneously calibrated on datasets reporting internal concentration in several organs in three fish species (original and literature datasets) by Bayesian methods (Monte Carlo Markov Chain). Model predictions were then compared to other experimental data (i.e., excluded from the calibration step) to assess the predictive performance of the model. The results strongly suggest that PCZ and IMZ are actively transported across the gills, resulting in a small fraction being effectively absorbed by the fish. The model's results also confirm that both molecules are extensively metabolized by the liver into mainly glucuronate conjugates. Overall, the model performances were satisfying, predicting internal concentrations in several key organs. On average, 90% of experimental data were predicted within a two-fold range. The PBK model allows the understanding of IMZ and PCZ kinetics profiles by accurately predicting internal concentrations in three different fish species regardless of the exposure scenario. This enables a proper understanding of the mechanism of action of EDs at the molecular initiating event (MIE) by predicting bioaccumulation in target organs, thus linking this MIE to a possible adverse outcome.

Bayesian ridge regression shows the best fit for SSR markers in Psidium guajava among Bayesian models.

Markers are an important tool in plant breeding, which can improve conventional phenotypic breeding, generating more accurate information outcoming better decision making. This study aimed to apply and compare the fit of different Bayesian models BRR, BayesA, BayesB, BayesB (setting the value from very low to [Formula: see text] = [Formula: see text]), BayesC and Bayesian Lasso (LASSO) for predictions of the genomic genetic values of productivity and quality traits of a guava population. The models were fitted for traits fruit mass, pulp mass, soluble solids content, fruit number, and production per plant in the genomic prediction with SSR markers, obtained through the CTAB extraction method with 200 primers. The Bayesian ridge regression model showed the best results for all traits and was chosen to predict the individual's genomic values according to the cross-validation data. A good stabilization of the Markov and Monte Carlo chains was observed with the mean values close to the observed phenotypic means. Heritabilities showed good predictive accuracy. The model showed strong correlations between some traits, allowing indirect selection.

A comparison of behavioral and reproductive parameters between wild-type, transgenic and mutant zebrafish: Could they all be considered the same "zebrafish" for reglementary assays on endocrine disruption?

Transgenic zebrafish models are efficiently used to study the effects of endocrine disrupting chemicals (EDC); thereby informing on their mechanisms of action. However, given the reported differences between zebrafish strains at the genetical, physiological and behavioral levels; care should be taken before using these transgenic models for EDC testing. In the present study, we undertook a set of experiments in different transgenic and/or mutant zebrafish strains of interest for EDC testing: casper, cyp19a1a-eGFP, cyp19a1a-eGFP-casper, cyp11c1-eGFP, cyp11c1-eGFP-casper. Some behavioral traits, and some biochemical and reproductive physiological endpoints commonly used in EDC testing were assessed and compared to those obtained in WT AB zebrafish to ensure that transgene insertion and/or mutations do not negatively modify basal reproductive physiology or behavior of the fish. Behavioral traits considered as anxiety and sociality have been monitored. Sociality was evaluated by monitoring the time spent near congeners in a shuttle box while anxiety was evaluated using the Novel tank diving test. No critical difference was observed between strains for either sociality or anxiety level. Concerning reproduction, no significant difference in the number of eggs laid per female, in the viability of eggs or in the female circulating VTG concentrations was noted between the 5 transgenic/mutants and the WT AB zebrafish studied. In summary, the transgene insertion and the mutations had no influence on the endpoints measured in basal conditions. These results were a prerequisite to the use of these transgenic/mutant models for EDC testing. Next step will be to determine the sensitivity of these biological models to chemical exposure to accurately validate their use in existing fish assays for EDC testing.

Refinement of an OECD test guideline for evaluating the effects of endocrine disrupting chemicals on aromatase gene expression and reproduction using novel transgenic cyp19a1a-eGFP zebrafish.

Transgenic fish are powerful models that can provide mechanistic information regarding the endocrine activity of test chemicals. In this study, our objective was to use a newly developed transgenic zebrafish line expressing eGFP under the control of the cyp19a1a promoter in the OECD Fish Short Term Reproduction Assay (TG 229) to provide additional mechanistic information on tested substances. For this purpose, we exposed adult transgenic zebrafish to a reference substance of the TG 229, i.e. prochloraz (PCZ; 1.7, 17.2 and 172.6 µg/L). In addition to "classical" endpoints used in the TG 229 (reproductive outputs, vitellogenin), the fluorescence intensity of the ovaries was monitored at 4 different times of exposure using in vivo imaging. Our data revealed that 172.6 µg/L PCZ significantly decreased the number of eggs laid per female per day and the concentrations of vitellogenin in females, reflecting the decreasing E2 synthesis due to the inhibition of the ovarian aromatase activities. At 7 and 14 days, GFP intensities in ovaries were similar over the treatment groups but significantly increased after 21 days at 17.2 and 172.6 µg/L. A similar profile was observed for the endogenous cyp19a1a expression measured by qPCR thereby confirming the reliability of the GFP measurement for assessing aromatase gene expression. The overexpression of the cyp19a1a gene likely reflects a compensatory response to the inhibitory action of PCZ on aromatase enzymatic activities. Overall, this study illustrates the feasibility of using the cyp19a1a-eGFP transgenic line for assessing the effect of PCZ in an OECD test guideline while providing complementary information on the time- and concentration-dependent effects of the compound, without disturbing reproduction of fish. The acquisition of this additional mechanistic information on a key target gene through in vivo fluorescence imaging of the ovaries was realized without increasing the number of individuals.