2019-12-23, Willi, Raphael, Fent, Karl, Schmid, Simon

2019-10-15, Christen, Verena, Krebs, Jana, Bünter, Ivan, Fent, Karl

2019-07-01, Willi, Raffael Alois, Faltermann, Susanne, Hettich, Timm, Fent, Karl

Many glucocorticoids occur in the aquatic environments but their adverse effects to fish are poorly known. Here we investigate effects of the natural glucocorticoid corticosterone and the synthetic glucocorticoids betamethasone and flumethasone in zebrafish embryos. Besides studying the effects of each steroid, we compared effects of natural with synthetic glucocorticoids, used as drugs. Exposure at concentrations of 1 μg/L and higher led to concentration-related decrease in spontaneous muscle contractions at 24 h post fertilization (hpf) and increase in heart rate at 48 hpf. Betamethasone showed a significant increase at 0.11 μg/L in heart rate. Corticosterone also accelerated hatching at 60 hpf at 0.085 μg/L. Transcription of up to 24 genes associated with different pathways showed alterations at 96 and 120 hpf for all glucocorticoids, although with low potency. Corticosterone caused transcriptional induction of interleukin-17, while betamethasone caused transcriptional down-regulation of the androgen receptor, aromatase and hsd11b2, indicating an effect on the sex hormone system. Furthermore, transcripts encoding proteins related to immune system regulation (irg1l, gilz) and fkbp5 were differentially expressed by corticosterone and betamethasone, while flumethasone caused only little effects, mainly alteration of the irg1l transcript. Our study shows that these glucocorticoids caused more potent physiological effects in early embryos than transcriptional alterations in hatched embryos, likely due to increased metabolism in later developmental stages. Thus, these glucocorticoids may be of concern for early stages of fish embryos in contaminated aquatic environments.

2019-01, Christen, Verena, Joho, Yvonne, Vogel, Maren, Fent, Karl

The pyrethroid deltamethrin and the organophosphate insecticide dimethoate are widely used in agriculture and in urban areas. Both plant protection products (PPPs) unintendedly result in adverse effects in pollinators. Currently, the sublethal effects of both compounds are poorly known, particularly on the molecular and biochemical level. Here we analysed effects of deltamethrin and dimethoate at environmental and sublethal concentrations in honey bee workers by focusing on transcriptional changes of target genes in the brain. In addition, expression of vitellogenin protein and activity of acetylcholinesterase were assessed upon dimethoate exposure to assess physiological effects. Deltamethrin resulted in induction of the cyp9q2 transcript at 0.53 ng/bee, while dimethoate led to induction of vitellogenin on the mRNA and protein level at 2 ng/bee. Transcripts of additional cytochrome P450-dependent monooxygenases (cyps) and genes related to immune system regulation were not differentially expressed upon PPP exposure. Dimethoate but not deltamethrin led to a strong and concentration-related inhibition of the acetylcholinesterase at 2 and 20 ng/bee. Our data demonstrate that deltamethrin and dimethoate exhibit transcriptional effects at environmental concentrations in the brain of honey bees. Dimethoate also strongly affected physiological traits, which may translate to adverse effects in forager bees.

2019-12-12, Willi, Raphael, Furia, Nathan, Fent, Karl, Faltermann, Susanne, Mastroianni, Sarah

2019-09-05, Christen, Verena, Krebs, Jana, Fent, Karl

Fungicides are highly used for plant protection but their molecular and chronic effects are poorly known. Here, we analyse transcriptional effects in the brain of honey bees of three frequently applied fungicides, azoxystrobin, chlorothanolin and folpet, after oral exposure for 24, 48 and 72 h. Among transcripts assessed were genes encoding proteins for immune and hormone system regulation, oxidative phosphorylation, metabolism, and acetylcholine receptor alpha 1. Azoxystrobin and folpet induced minor alterations, including down-regulation of hbg-3 by azoxystrobin and induction of ndufb-7 by folpet. Chlorothanolin induced strong transcriptional down-regulation of genes encoding enzymes related to oxidative phosphorylation and metabolism, including cyp9q1, cyp9q2 and cyp9q3, acetylcholine receptor alpha 1 and hbg-3 and ilp-1, which are linked to hormonal regulation and behavioural transition of honey bees. Exposures to chlorothanolin in different seasonal times showed different responsiveness; responses were faster and often stronger in April than in June. Chlorothanolin caused the strongest effects and affected transcriptional abundance of genes related to energy production, metabolism and the endocrine system. Disturbed energy production may reduce foraging activity and hormonal dysregulation, such as the transition of nurse bees to foragers. Further analyses are needed to further substantiate potential adverse effects of chlorothanolin in bees on the physiological level.

2019-02, Christen, Verena, Fent, Karl, Hettich, Timm

The insect yolk precursor vitellogenin is a lipoglycoprotein synthesized and stored in the fat body and secreted into the hemolymph. In honey bees, vitellogenin displays crucial functions in hormone signaling, behavioral transition of nurse bees to foragers, stress resistance, and longevity in workers. Plant protection products such as neonicotinoids, pyrethroids, and organophosphates alter the transcriptional expression of vitellogenin. To assess plant protection product‐induced alterations on the protein level, we developed a rabbit polyclonal vitellogenin antibody. After characterization, we assessed its specificity and vitellogenin levels in different tissues of worker bees. The vitellogenin antibody recognized full‐length 180‐kDa vitellogenin and the lighter fragment of 150 kDa in fat body, hemolymph, and brain. In hemolymph, a band of approximately 75 kDa was detected. Subsequent mass spectrometric analysis (liquid chromatography–mass spectrometry) confirmed the 180‐ and 150‐kDa bands as vitellogenin. Subsequently, we evaluated vitellogenin expression in brain, fat body, and hemolymph on 24‐h exposure of bees to 3 ng/bee to the neonicotinoid clothianidin. Full‐length vitellogenin was upregulated 3‐fold in the fat body, and the 150‐kDa fragment was upregulated in the brain of exposed honey bees, whereas no alteration occurred in the hemolymph. Upregulation of the vitellogenin protein by the neonicotinoid clothianidin is in line with the previously shown induction of its transcript. We conclude that vitellogenin might serve as a potential biomarker for neonicotinoid and other pesticide exposure in bees. Environ Toxicol Chem 2019;00:1–10. © 2019 SETAC

2019-12-04, Fent, Karl, Christen, Verena

2019-07-15, Willi, Raffael Alois, Carcaiso, Giulia, Fent, Karl

Many synthetic glucocorticoids from medical applications occur in the aquatic environment. Whether they pose a risk for fish health is poorly known. Here we investigate effects of glucocorticoids fluticasone propionate (FLU) and triamcinolone acetonide (TRI) as single steroids and as ternary mixtures with clobetasol propionate (CLO) in zebrafish embryos. Exposure to FLU and TRI in a range of concentrations between 0.099 and 120.08 μg/L led to concentration-related decrease in muscle contractions and increase in heart rate at 0.98 and 1.05 μg/L, respectively, and higher. Genes encoding for proteins related to glucose metabolism (g6pca, pepck1), immune system regulation (fkbp5, irg1l, socs3, gilz) and matrix metalloproteinases mmp-9 and mmp-13 showed expressional alterations, as well as genes encoding for the progestin receptor (pgr) and corticosteroid dehydrogenase (hsd11b2). FLU accelerated hatching and led to embryotoxicity (immobilization and edema). Ternary mixtures (FLU + TRI + CLO) induced the same physiological and toxicological effects at concentrations of individual glucocorticoids of 11.1–16.37 μg/L and higher. Heart rate was increased in the mixture at concentrations as low as 0.0885−0.11 μg/L of each steroid. Glucocorticoids in mixtures showed additive activity; the fold-changes of transcripts of 19 target genes were additive. Together, our data show that glucocorticoids act additively and their joint activity may be of concern for developing fish in contaminated environments.

2019-01, Brun, Nadja, Fent, Karl

Aquatic systems are contaminated by many metals but their effects as mixtures on organisms are not well understood. Here, we assessed effects of aluminum with fairly well-known modes of actions and indium, an understudied emerging contaminant from electronics, followed by studying equi-effective mixtures thereof. We report acute and adverse phenotypic effects in Daphnia magna adults and global transcriptomic effects employing RNA sequencing in neonates. The mixture induced more than additive activity in mortality and in physiological effects, including growth and reproduction. Similarly, transcriptomic effects were more than additive, as indicated by a markedly higher number of 463 differentially expressed transcripts in the mixture and by distinct classes of genes assigned to several biological functions, including metabolic processes, suggesting depleted energy reserves, which may be responsible for the observed impaired reproduction and growth. A gene set enrichment analysis (GSEA) of a priori known response pathways for aluminum confirmed activation of distinct molecular pathways by indium. Our study is highlighting more than additive effects at the transcriptional and physiological level and is providing a state-of-the art approach to mixture analysis, which is important for risk assessment of these metals and metal mixtures.