Effects of exposure to 17-alpha-ethynylestradiol on sperm quality of tench (Tinca tinca).

Alterations of sperm quality were studied in tench (Tinca tinca) exposed to sub-lethal doses of 17-alpha-ethynylestradiol-EE2-(50, 100 and 500µg/kg t.w) under semi-static conditions for 30 days. Thus, different biomarkers of sperm quality were assessed: concentration and volume of ejaculate, total number of spermatozoa, percentage of motile spermatozoa, sperm motility and percentage of live and dead spermatozoa. Sperm motility was examined by computer-assisted image analysis and the viability of spermatozoa was assessed through flow cytometry. The most relevant alterations observed were significant reductions in the reproductive parameters such as testicular somatic index, spermatozoa concentration, straight line velocity, curvilinear velocity, average path velocity and wobble in tench exposed to 50µg/kg t.w of EE2. Our study about the effects of EE2 on the sperm quality in tench provides new evidences which strengthen the fact that this synthetic estrogen is included in the list of non-monotonic dose response compounds in animal studies.

Histological alterations in the structure of the testis in tench (Tinca tinca) after exposure to 17 alpha-ethynylestradiol.

Environmental pollution with synthetic estrogens may pose a serious threat to reproduction of aquatic wildlife species. The current study describes the effects of 17α-ethynylestradiol (EE2 ) on the structure of the testis in tench (Tinca tinca). Adult male tench were exposed to sublethal doses of EE2 (50, 100, and 500 µg/Kg t.w.) under semistatic conditions for a period of 30 days. The condition factor (CF), testicular somatic index (TSI), and histology (including a morphometric analysis) of the testis were examined. No consistent differences were observed in the CF of EE2 -exposed tench when compared with nonexposed fish. A significant decrease in TSI could only be observed at a 50 µg/Kg t.w. EE2 dose (p < 0.05) when compared with the control group. The histopathology of the testis was associated with loss of normal tubular structure with increased doses of exposure, decrease of tubule number, degeneration in Sertoli and Leydig cells, increase in necrotic testicular cells including formation of syncytia structures and, finally, a high incidence of fish with early primary oocytes at 100 and 500 µg/Kg t.w. EE2 . These results indicate that long-term exposure to EE2 may produce clear negative effects on testicular structure in tench.

Histological alterations on the structure of the excretory renal system in tench (Tinca tinca) after exposure to 17-alpha-ethynylestradiol.

This study describes the effects of 17-alpha-ethynylestradiol (EE2) on the structure of the excretory system of the kidney in tench. Adult male tench were exposed to sub-lethal doses of EE2 (50, 100 and 500 µg/kg b.w.) under semistatic conditions for a period of 30 days. The nephrosomatic index and histology (including a morphometric analysis) of the kidney were examined. Histopathological lesions in the kidney of exposed tench were: dilation of glomerular capillaries and increase in the area of the renal corpuscle, hyaline degeneration in the epithelial cells of the proximal tubules leading to necrotic changes, hemorrhages in the interstitial tissue and deposits of eosinophilic material. These lesions were observed with a greater degree of severity as the exposure doses were increased. These results indicate that long-term exposure to EE2 could produce clear negative effects on the excretory system of the kidney in tench and consequently on their physiological functions.

Effect of long-term exposure to simazine on histopathology, hematological, and biochemical parameters in Cyprinus carpio.

We investigated if residues of simazine in the natural waters would cause histological, hematological, and biochemical alterations in carps from contaminated areas in Badajoz (Spain). Some necrotic foci in kidney and liver, hepatitis, and hepatic steatosis were detected. No changes on measured hematological and biochemical parameters between fish from reference and contaminated ponds were observed. To assess if simazine exposure was the cause of these observations carps were exposed in the laboratory to simazine (45 microg/L) for 90 days. Some results obtained in the field were confirmed in laboratory, such as necrosis in kidney and liver and hepatic steatosis. Globular eosinophilic foci in kidney and a slight decrease of the hematocrit were also detected. These changes were moderate and indicative of an adaptation of the fish to the toxic stress caused by exposure to low simazine concentrations.

Assessment of the effects of the carbamazepine on the endogenous endocrine system of Daphnia magna.

In the present study, the endocrine activity of the antiepileptic pharmaceutical carbamazepine (CBZ) in the crustacean Daphnia magna was assessed. To assess the hormonal activity of the drug, we exposed maternal daphnids and embryos to environmental relevant concentrations of CBZ (ranging from 10 to 200 µg/L) and to mixtures of CBZ with fenoxycarb (FEN; 1 µg/L). Chronic exposure to CBZ significantly decreased the reproductive output and the number of molts of D. magna at 200 µg/L. This compound induced the production of male offspring (12 ± 1.7 %), in a non-concentration-dependent manner, acting as a weak juvenile hormone analog. Results showed that this substance, at tested concentrations, did not antagonize the juvenoid action of FEN. Further, CBZ has shown to be toxic to daphnid embryos through maternal exposure interfering with their normal gastrulation and organogenesis stages but not producing direct embryo toxicity. These findings suggest that CBZ could act as an endocrine disruptor in D. magna as it decreases the reproductive output, interferes with sex determination, and causes development abnormality in offspring. Therefore, CBZ could directly affect the population sustainability.

A new Bayesian network-based approach to the analysis of sperm motility: application in the study of tench (Tinca tinca) semen.

In this study a Bayesian network (BN) has been built for the study of the objective motility of Tinca tinca spermatozoa (spz). Semen from eight 2-year-old sexually mature male tenchs was obtained and motility analyses were performed at 6-17, 23-34 and 40-51 s after activation, using computer-assisted sperm analysis (CASA) software. Motility parameters rendered by CASA were treated with a two-step cluster analysis. Three well-defined sperm subpopulations were identified, varying the proportion of spermatozoa contained in each cluster with time and male. Cluster, cinematic and time variables were used to build the BN to study the probabilistic relationships among variables and how each variable influenced the final sperm classification into one of three predefined clusters. Both network structure and conditional probabilities were calculated based on the collected data set. Results shown that almost all the variables were directly or indirectly related to each other. By doing probabilistic inference we observed that the cluster distribution corresponded to the definition provided by the cluster analysis. Also, velocity and time variables determined the cluster to which each spermatozoon belonged with a high degree of accuracy. Thus, BNs can be applied in the study of sperm motility. The construction of a BN that include fertility data opens a new way to try to clarify the roles of motility and other sperm quality indicators in fertilization.

Effect of long-term exposure to simazine on brain and muscle acetylcholinesterase activity of common carp (Cyprinus carpio).

Several water-contamination incidents with simazine have occurred in the province of Badajoz (Spain), due to its excessive use for controlling weeds in olive trees and vineyards. Simazine residues were also detected in drinking water, increasing public health concern. However, little is known on the effects that low levels of simazine pose to environment organisms. We investigated if residues of simazine in the natural waters would affect brain and muscle acetylcholinesterase activity in common carps captured in areas in which simazine residues were detected at average levels of 4.5 microg/L. Results confirmed depression on brain and muscle acetylcholinesterase activity of 20% and 29%, respectively, in carps inhabiting one of the simazine-contaminated ponds, termed "Molinos de Matachel." To assess the biological significance of this finding, we developed a controlled laboratory study in which carps were exposed to simazine at 45 microg/L (10-fold that of the natural water levels) for 90 days. The results obtained in the field study were not confirmed in our laboratory experiment, since carps did not show evidence or brain or muscle acetylcholinesterase activity depression for the duration of the experiment, and therefore, we can conclude that acetylcholinesterase depression found in carps collected in "Molinos de Matachel" should be ascribed to other compounds or mixtures of xenobiotics.