Oxidative stress, biotransformation enzymes and histopathological alterations in Nile tilapia (Oreochromis niloticus) exposed to new and used automotive lubricant oil.

Lubricant oils are among oil-based products that are not fully consumed during its use, thereby producing non-biodegradable residues which can cause contamination of natural systems. This study evaluated the toxicity of new and used lubricating oil (0.01 and 0.1 mL L-1) in adult Nile tilapia (Oreochromis niloticus), by assessing the effects on oxidative stress, biotransformation enzymes (liver and gills), and histopathological alterations on hepatic and pancreatic tissues after 3 and 7 days of exposure. Results showed that 3-days exposure to 0.1 mL L-1 of used and new lubricating oil increased the activity of superoxide dismutase (SOD) and malondialdehyde (MDA) levels in liver of O. niloticus, respectively. In gills, catalase (CAT) was decreased in fish exposed to 0.1 mL L-1 of non-used oil after 3 days, but pronounced increases in CAT was detected after 7 days-exposure to both new and used oil. Shorter exposure to both concentrations of new and used oil also raised glutathione-S-transferase activity (GST) in gills. Ethoxyresorufin-O-deethylase (EROD) was induced in liver of fish exposed to 0.1 mL L-1of used oil after 3 and 7 days, however a reduced response of this enzyme was detected in gills of animals from both oil treatments. In vitro analysis showed that hepatic EROD was inhibited by lubricating oil exposures, with more pronounced responses in treatments containing used oil. Hepatic lesions, such as cytoplasmic vacuolization, nuclei abnormally, changes in hepatocytes shape, steatosis, cholestasis, eosinophilic inclusions and necrosis were mainly increased by 7 days exposure to used lubricating oil at higher concentration.

Effects of methylphenidate on the aggressive behavior, serotonin and dopamine levels, and dopamine-related gene transcription in brain of male Nile tilapia (Oreochromis niloticus).

The occurrence of pharmaceuticals in the aquatic environment has increased considerably in the last decades, causing negative biochemical, physiological, and behavioral effects in aquatic organisms. In this study, we evaluated the effects of methylphenidate (MPH) on the aggressive behavior, dopamine-related gene transcript levels, monoamine levels, and carboxylesterase transcript levels and activity in the brain of male Nile tilapia (Oreochromis niloticus). Carboxylesterase activity was also measured in the liver and gills. Fish were exposed for 5 days to MPH at 20 and 100 ng L-1. Fish exposed to 100 ng L-1 of MPH showed increased aggressiveness and decreased dopamine (DA) and serotonin (5-HT) levels. No changes were observed in plasma testosterone levels and in the transcript levels of D1 and D2 dopamine receptors, dopamine transporter (DAT), and carboxylesterase 2 (CES2). Exposure to 100 ng L-1 of MPH caused a decrease in the transcript levels of carboxylesterase 3 (CES3) and an increase in tyrosine hydroxylase (TH), while exposure to 20 ng L-1 of MPH increased the transcript levels of D5 dopamine receptor. Carboxylesterase activity was unchanged in the brain and liver and increased in the gills of fish exposed to 20 ng L-1. These results indicate that MPH at 100 ng L-1 increases aggressiveness in Nile tilapia, possibly due to a decrease in 5-HT levels in the brain and alterations in dopamine levels and dopamine-related genes.

Estrogenic and anti-androgenic effects of the herbicide tebuthiuron in male Nile tilapia (Oreochromis niloticus).

Tebuthiuron is a phenylurea herbicide widely used in agriculture that can reach the aquatic environments, possibly posing negative effects to the aquatic biota. Phenylurea herbicides, such as diuron, are known to cause estrogenic and anti-androgenic effects in fish, but no such effects were yet reported for tebuthiuron exposure. Thus, the aim of this study was to evaluate if tebuthiuron, at environmentally relevant concentrations (100 and 200ng/L) and after 25days of exposure have estrogenic and/or anti-androgenic effects on male of Nile tilapia (Oreochromis niloticus), through the evaluation of plasmatic testosterone (T) and estradiol (E2) levels, brain aromatase (CYP19) levels (western-blot), and by evaluating the histology of the testicles. When compared to the control group, plasmatic T levels decreased about 76% in the animals exposed to 200ng/L of tebuthiuron, while E2 levels increased about 94%, which could be related to a significant increase (77%) in CYP19A1 levels, an enzyme that catalyzes the conversion of androgens into estrogens. Histological analyses of the testicles also demonstrated that tebuthiuron at both tested concentrations caused a decrease in the diameter of the seminiferous tubules and in the diameter of the lumen. Therefore, the gonadosomatic index (GSI) was reduced by 36% % in the animals exposed 200ng/L to tebuthiuron. Indeed, the relative frequency of spermatocytes and spermatids increased respectively 73% (200ng/L) and 61% (100ng/L) in the tebuthiuron exposed animals, possibly due to the impairment of sperm release into the lumen, that was decreased 93% (200ng/L) in the treated animals compared to the control. These results confirm that tebuthiuron causes estrogenic and anti-androgenic effects in Nile tilapias at environmentally relevant concentrations.

Diuron metabolites act as endocrine disruptors and alter aggressive behavior in Nile tilapia (Oreochromis niloticus).

Diuron and its biodegradation metabolites were recently reported to cause alterations in plasma steroid hormone concentrations with subsequent impacts on reproductive development in fish. Since steroid hormone biosynthesis is regulated through neurotransmission of the central nervous system (CNS), studies were conducted to determine whether neurotransmitters that control hormone biosynthesis could be affected after diuron and diuron metabolites treatment. As the same neurotransmitters and steroid hormones regulate behavioral outcomes, aggression was also evaluated in male Nile tilapia (Oreochromis niloticus). Male tilapias were exposed for 10 days to waterborne diuron and the metabolites 3,4-dichloroaniline (DCA), 3,4-dichlorophenyl-N-methylurea (DCPMU), at nominal concentrations of 100 ng L-1. In contrast to Diuron, DCA and DCPMU significantly diminished plasma testosterone concentrations (39.4% and 36.8%, respectively) and reduced dopamine levels in the brain (47.1% and 44.2%, respectively). In addition, concentrations of the stress steroid, cortisol were increased after DCA (71.0%) and DCPMU (57.8-%) exposure. A significant decrease in aggressive behavior was also observed in animals treated with the metabolites DCA (50.9%) and DCPMU (68.8%). These results indicate that biotransformation of diuron to active metabolites alter signaling pathways of the CNS which may impact androgen and the stress response as well as behavior necessary for social dominance, growth, and reproduction.

Leporinus elongatus induced spawning using carp pituitary extract or mammalian GnRH analogue combined with dopamine receptor antagonists.

Several studies have been developed to support the replacement of the crude carp pituitary extract (CPE) by synthetic products for induced reproduction of South American rheophilic species. However, results have been quite heterogeneous and there is no consensus or a routine use of synthetic products in these species. Thus, the aim of this study was to evaluate the ovulatory process in L. elongatus using different protocols of hormonal induction. Thus, fifteen wild mature females maintained at the Experimental Fish Station, Salto Grande, SP, Brazil were submitted to three different hormonal treatments: CPE (fractioned dose: 0.5 and 5.0 mg kg-1); mGnRHa (single dose: 3.5 µg kg-1) and mGnRHa (single dose: 5.0 µg kg-1). The spawning rate and absolute fecundity were similar among the treatments, but fertility rates were higher for CPE treatment (23.60 ± 9.40) then for mGnRHa treatments (close to or zero zero). Although females ovulated in all treatments, none of them provided viable embryos, showing hatching rates close to zero or zero. Both mGnRHa treatments were more potent for inducing the ovulatory process then CPE treatment, which was evidenced by the fact that the formers showed higher volume density of postovulatory follicles (POF). Accordingly, E2 and 17α-OHP plasma levels were higher for the mGnRHa treated females compared to the CPE one at the time of ovulation. In this study we confirmed previous scientific evidence that, regardless of whether promoting ovulation, the use of conventional CPE and GnRH doses are not appropriate for some South American migratory species, due to the non-attainment of viable embryos. Moreover, we have brought new information about the relationship between reproductive performance and gonadal steroids concentrations using different hormonal therapies, contributing to understand the reasons for Leporinus elongatus embryo loss in induced spawning.

Estrogenic activities of diuron metabolites in female Nile tilapia (Oreochromis niloticus).

Some endocrine disrupting chemicals (EDCs) can alter the estrogenic activities of the organism by directly interacting with estrogen receptors (ER) or indirectly through the hypothalamus-pituitary-gonadal axis. Recent studies in male Nile tilapia (Oreochromis niloticus) indicated that diuron may have anti-androgenic activity augmented by biotransformation. In this study, the effects of diuron and three of its metabolites were evaluated in female tilapia. Sexually mature female fish were exposed for 25 days to diuron, as well as to its metabolites 3,4-dichloroaniline (DCA), 3,4-dichlorophenylurea (DCPU) and 3,4-dichlorophenyl-N-methylurea (DCPMU), at concentrations of 100 ng/L. Diuron metabolites caused increases in E2 plasma levels, gonadosomatic indices and in the percentage of final vitellogenic oocytes. Moreover, diuron and its metabolites caused a decrease in germinative cells. Significant differences in plasma concentrations of the estrogen precursor and gonadal regulator17α-hydroxyprogesterone (17α-OHP) were not observed. These results show that diuron metabolites had estrogenic effects potentially mediated through enhanced estradiol biosynthesis and accelerated the ovarian development of O. niloticus females.

Anti-androgenic activities of diuron and its metabolites in male Nile tilapia (Oreochromis niloticus).

Diuron (3-(3,4-dichlorophenyl)-1,1-dimethylurea) is a widely used herbicide which has been frequently detected in surface waters throughout the world. In vivo bioassay guided fractionation studies indicated that diuron may have estrogenic activity augmented by biotransformation. This study evaluated the effects of diuron and three of its metabolites on plasma hormone concentrations and spermatogenesis of the freshwater fish Nile tilapia (Oreochromis niloticus). Sexually mature male fish were exposed for 25 days to diuron, as well to its metabolites 3,4-dichloroaniline (DCA), 3,4-dichlorophenylurea (DCPU) and 3,4-dichlorophenyl-N-methylurea (DCPMU), at concentrations of 200ng/L. Testosterone levels were decreased by diuron, but had limited effects on gonadal histology. Diuron metabolites, however, caused significant decreases in testosterone and in 11-ketotestosterone, gonadosomatic index, diameter of seminiferous tubules and in the mean percentages of germ cells (spermatids and spermatozoa). We conclude that these metabolites have antiandrogenic activity to male Nile tilapia, potentially causing reproductive impairment in male fish.