Plasma Vitellogenin Research Articles

Biological effects and bioaccumulation of steroidal and phenolic endocrine disrupting chemicals in high-back crucian carp exposed to wastewater treatment plant effluents

Endocrine disrupting chemicals (EDCs) found in wastewater treatment plant (WWTP) effluents have been shown to cause adverse effects, but the uptake of EDCs from effluents (measured in fish muscle) are not known. In this study, the biological effects and bioaccumulation of steroidal and phenolic EDCs were assessed in high-back crucian carp ( Carassius auratus) exposed to WWTP effluents for 141 days. Compared with fish controls caged in Dianchi Lake, a significant reduction in gonadosomatic index (GSI) and increase in hepatosomatic index (HSI) and plasma vitellogenin (VTG) levels were observed in effluent-exposed fish. The concentrations of steroids and phenols in effluent-exposed fish showed time-dependent increase during the exposure. In addition, bioconcentration factors (BCFs) for steroids and phenols were between 17 and 59 on day 141. The results confirm that steroids and phenols bioconcentrate in fish muscle and this accumulation may account for the biological effects associated with exposures to WWTP effluents.

PCR–ELISA detection of estrogen receptor β mRNA expression and plasma vitellogenin induction in juvenile sole (Solea solea) exposed to waterborne 4-nonylphenol

In this study, the effect of 4-nonylphenol (4NP) on reproductive axis of sole (Solea solea) has been investigated by using selected biomarkers of estrogenic effects: vitellogenin (VTG) and estrogen receptor β (ERβ) mRNA. Furthermore, an enzyme linked immunosorbent assay (ELISA) detection system of reverse transcriptase-polymerase chain reaction (RT-PCR) products for the analysis of sole ERβ mRNA expression was developed and validated. The proposed method allows large-scale analyses of different mRNAs in fast and not expensive way. Our results have demonstrated that the PCR–ELISA method developed shows high sensitivity, good reproducibility and also the potential for semi-quantitative analysis of hepatic ERβ mRNAs. Both plasma VTG level and ERβ mRNA expression were increased in tested animals following a short exposure to environmental relevant concentrations (10−6M) of 4NP, confirming the functional role of ERβ in the regulation of xenoestrogens-induced production of VTG. The methodology provided in the present study together with the preliminary results on the hepatic expression of ERβ may be useful in environmental xenoestrogens monitoring studies, using flatfish as “sentinel” species.

Assessing the Effects of Historical Exposure to Endocrine-Active Compounds on Reproductive Health and Genetic Diversity in Walleye, a Native Apex Predator, in a Large Riverine System

In this combined field and laboratory study, we assessed whether populations of native walleye in the Upper Mississippi River experienced altered genetic diversity correlated with exposure to estrogenic endocrine-active compounds (EACs). We collected fin-clips for genetic analysis from almost 600 walleye (13 sites) and subsampled 377 of these fish (6 sites) for blood and reproductive organs. Finally, we caged male fathead minnows at 5 sampling sites to confirm the presence of estrogenic EACs. Our findings indicate that male walleye in four river segments produced measurable concentrations of plasma vitellogenin (an egg-yolk protein and, when expressed in male fish, a biomarker of acute estrogenic exposure), a finding consistent with the presence of estrogenic EACs and consistent with published historical data for at least three of these study sites (Grand Rapids, St. Paul, and Lake City on Lake Pepin). Patterns of vitellogenin induction were consistent for native walleye and caged fathead minnows. No widespread occurrence of histopathological changes, such as intersex was found compared with published reports of intersex at the furthest downstream study site. To assess possible effects of estrogenic exposure on the genetic diversity of walleye populations at the study sites, we DNA-fingerprinted individual fish using 10 microsatellite loci. Genetic differences were observed between populations; however, these differences were consistent with geographic distance between populations, with the largest observed difference in genetic diversity found between fish upstream and downstream of St. Anthony Falls (and/or Lock and Dam 1 of the Mississippi River), traditionally a historical barrier to upstream fish movement. Although the persistent occurrence of endocrine disruption in wild fish populations is troubling, we did not detect degradation of reproductive organs in individual walleye or alteration in genetic diversity of walleye populations.

Biomarker responses in Atlantic cod (Gadus morhua) exposed to produced water from a North Sea oil field: Laboratory and field assessments

Biological markers of produced water (PW) exposure were studied in Atlantic cod (Gadus morhua) in both laboratory and field experiments, using authentic PW from a North Sea oil field. In the laboratory study, the PW exposure yielded significantly elevated levels of metabolites of polycyclic aromatic hydrocarbons (PAHs) and alkylphenols (APs) in bile even at the lowest exposure dose (0.125% PW). Other biomarkers (hepatic CYP1A induction and DNA adduct formation) responded at 0.25% and 0.5% PW concentrations. In the field study, bile metabolite markers and hepatic CYP1A were clearly increased in fish caged close to the PW outfall. Induction of plasma vitellogenin was not found in laboratory or field exposures, suggesting that the levels of oestrogen agonists (such as APs) might not have been sufficient to elicit induction, under the present conditions. The applicability of the biomarkers for use in water column biomonitoring programs is discussed.

Characterization of Plasma Vitellogenin and Sex Hormone Concentrations during the Annual Reproductive Cycle of the Endangered Razorback Sucker

AbstractPopulation declines of the endangered razorback sucker Xyrauchen texanus in the Colorado River basin have been attributed to predation by and competition with nonnative fishes, habitat alteration, and dam construction. The reproductive health and seasonal variation of the reproductive end points of razorback sucker populations are currently unknown. Using nonlethal methods, we characterized the plasma hormonal fluctuations of reproductively mature female and male razorback suckers over a 12‐month period in a hatchery by measuring their vitellogenin (VTG) and three sex hormones: 17β‐estradiol (E2), testosterone (T), and 11‐ketotestosterone (KT). Fish were identified as reproductive or nonreproductive based on their body weight, VTG, and sex hormone profiles. In reproductive females, the E2 concentration increased in the fall and winter, and increases in T and VTG concentrations were generally associated with the spawning period. Mean T concentrations were consistently greater in reproductive females than in nonreproductive females, but this pattern was even more pronounced during the spawning period (spring). Consistently low T concentrations (<3 ng/mL) in adult females during the spawning period may indicate reproductive impairment. In reproductive males, spring increases in KT and T concentrations were associated with spawning; concentrations of E2 (<0.48 ng/mL) and VTG (<0.001 mg/mL) were low in males throughout the study. In addition, the E2 : KT ratio and T were the best metrics by which to distinguish female from male adult razorback suckers throughout the year. These metrics of reproductive health and condition may be particularly important to recovery efforts of razorback suckers given that the few remaining wild populations are located in a river where water quality and quantity issues are well documented. In addition to the size, age, and recruitment information currently considered in the recovery goals of this endangered species, reproductive end points could be included as recovery metrics with which to monitor seasonal trends and determine whether repatriated populations are cycling naturally.Received September 24, 2010; accepted February 18, 2011

Effects of butachlor on reproduction and hormone levels in adult zebrafish (Danio rerio)

Butachlor, a chloracetamide herbicide, is widely used in China. In the present study, paired adult male and female zebrafish (Danio rerio) were exposed to various concentrations of butachlor (0, 25, 50 and 100 μg/L) for 30 days, and the effects on reproduction and endocrine disruption were evaluated using fecundity, condition factor (CF), gonadosomatic index (GSI), liver somatic index (LSI), plasma vitellogenin (VTG), sex steroids and thyroid hormone levels as endpoints. Our results showed that the mean fecundity rates were significantly decreased at 50 and 100 μg/L butachlor during the 30-day exposure period. At the end of the exposure period, no significant changes were observed in CF and LSI in both females and males, while GSI was significantly reduced in males at 50 and 100 μg/L butachlor. At 100 μg/L butachlor, plasma testosterone (T) and 17β-estradiol (E2) levels were significantly decreased in females, while plasma VTG level was significantly increased in males. Plasma thyroxine (T4) and triiodothyronine (T3) levels were significantly increased at 50 and 100 μg/L butachlor in males, and at 100 μg/L in females. This work demonstrated that butachlor adversely affected the normal reproductive success of zebrafish, and disrupted the thyroid and sex steroid endocrine systems, which provides the basis for the estimated ecological risk during butachlor exposure.

Effects of β‐endosulfan on the growth and reproduction of zebrafish (Danio rerio)

Because of persistent organic pollution in aquatic environments, the widely used organochlorine pesticide endosulfan, which is a potential endocrine disruptor, is expected to pose a significant risk to aquatic organisms. In the present study, we explored the potential endocrine-disrupting risk of β-endosulfan by investigating its effect on the growth, reproduction, plasma vitellogenin, and organ histology of adult zebrafish. We found that, although β-endosulfan did not significantly affect the growth of zebrafish, it greatly decreased the hatching rate, even at a concentration as low as 10 ng/L. Interestingly, the decrease of the hatching rate was highly correlated with pathological alterations of the testes. Additionally, the values of the gonadosomatic index were significantly reduced in female zebrafish treated with 200 ng/L β-endosulfan, which was also closely associated with ovarian histological changes. More importantly, a significant increase in the level of vitellogenin was observed in all male fish treated with β-endosulfan. Based on these findings, we conclude that β-endosulfan severely affects the reproductive function of zebrafish and the synthesis of vitellogenin in the liver, and thus, β-endosulfan has a serious endocrine disruption function in zebrafish.

Uptake of 17β-estradiol and biomarker responses in brown trout ( Salmo trutta) exposed to pulses

In streams, chemicals such as 17β-estradiol (E2) are likely to occur in pulses. We investigated uptake and biomarker responses in juvenile brown trout ( Salmo trutta) of 3- or 6-h pulses of concentrations up to 370 ng E2 L −1. Uptake by the fish was estimated from disappearance of E2 from tank water. A single 6-h pulse of 370 ng E2 L −1 increased the plasma vitellogenin concentration, liver Erα- and vitellogenin-mRNA. Exposure to 150–160 ng E2 L −1 for 6 h increased vitellogenin in one experiment but not in another. Two 6-h pulses had a larger effect one pulse. Brown trout in the size range 24–74 g took up E2 linearly with time and exposure concentration with a concentration ratio rate of 20.2 h −1. In conclusion, the threshold for induction of estrogenic effects in juvenile brown trout at short term pulse exposure appears to be in the range 150–200 ng E2 L −1.

Comparing biological effects and potencies of estrone and 17β-estradiol in mature fathead minnows, Pimephales promelas

The presence of endocrine active compounds such as estrogens in treated wastewater effluent and their effects on aquatic life are causing concern among aquatic resource managers. In contrast to 17β-estradiol (E2), the steroid hormone produced by all vertebrates, the biological effects of estrone (E1), one of its breakdown products are less understood, even though the aquatic concentrations of E1 are often higher than those of E2. The central hypothesis of this study was that at environmental concentrations, E1 has estrogenic effects in fish, with increased vitellogenin concentrations and decreased reproductive success in both male and female fathead minnows, as found with E2. In two replicate experiments, we exposed mature fathead minnows to three concentrations of each estrogen for 21 days in a flow-through exposure system and measured a broad suite of anatomical (body indices, histopathology), physiological (plasma vitellogenin), behavioral (nest defense), and reproductive (fecundity, fertility, hatching) endpoints. These endpoints have previously been associated with adverse effects of estrogenic exposures. While body length and weight parameters were unaltered by exposure, secondary sex characteristics exhibited an exposure concentrated-related decline in male fathead minnows. Interestingly, low concentrations of estrone (∼15 ng/L) enhanced the aggressiveness of male fathead minnows in a behavioral assay. Vitellogenin concentrations in male fish increased with higher concentrations of both estrogens, but remained unchanged in all female treatments. A decrease in fecundity was observed at high concentrations of E2 as compared with control minnows. These results suggest that E1, at concentrations previously found in waters receiving wastewater effluent, can have reproductive effects on fish.

Reproductive endocrine disruption in smallmouth bass (Micropterus dolomieu) in the Potomac River basin: spatial and temporal comparisons of biological effects

A high prevalence of intersex or testicular oocytes (TO) in male smallmouth bass within the Potomac River drainage has raised concerns as to the health of the river. Studies were conducted to document biomarker responses both temporally and spatially to better understand the influence of normal physiological cycles, as well as water quality and land-use influences. Smallmouth bass were collected over a 2-year period from three tributaries of the Potomac River: the Shenandoah River, the South Branch Potomac and Conococheague Creek, and an out-of-basin reference site on the Gauley River. The prevalence of TO varied seasonally with the lowest prevalence observed in July, post-spawn. Reproductive maturity and/or lack of spawning the previous spring, as well as land-use practices such as application of manure and pesticides, may influence the seasonal observations. Annual, seasonal, and site differences were also observed in the percentage of males with measurable concentrations of plasma vitellogenin, mean concentration of plasma vitellogenin in females, and plasma concentrations of 17β-estradiol and testosterone in both sexes. Bass collected in the South Branch Potomac (moderate to high prevalence of TO) had less sperm per testes mass with a lower percentage of those sperm being motile when compared to those from the Gauley River (low prevalence of TO). An inverse relationship was noted between TO severity and sperm motility. An association between TO severity and wastewater treatment plant flow, percent of agriculture, total number of animal feeding operations, the number of poultry houses, and animal density within the catchment was observed.

The influence of 17β-estradiol on intestinal calcium carbonate precipitation and osmoregulation in seawater-acclimated rainbow trout (Oncorhynchus mykiss)

The intestine of marine teleosts produces carbonate precipitates from ingested calcium as part of their osmoregulatory strategy in seawater. The potential for estrogens to control the production of intestinal calcium carbonate and so influence osmoregulation was investigated in seawater-acclimated rainbow trout following intraperitoneal implantation of 17β-estradiol (E2) at two doses (0.1 and 10 μg E2 g(-1)). Levels of plasma vitellogenin provided an indicator of estrogenic effect, increasing significantly by three and four orders of magnitude at the low and high doses, respectively. Plasma osmolality and muscle water content were unaffected, whereas E2-treated fish maintained lower plasma [Na(+)] and [Cl(-)]. Plasma [Ca(2+)] and [Mg(2+)] and muscle [Ca(2+)] increased with vitellogenin induction, whereas the intestinal excretion of calcium carbonate was reduced. This suggests that elevated levels of circulating E2 may enhance Ca(2+) uptake via the gut and simultaneously reduce CaCO(3) formation, which normally limits intestinal availability of Ca(2+). Increasing E2 caused an elevation of [Na(+)] and [Cl(-)] and a reduction of [HCO(3(-))] in intestinal fluid. We speculate that E2 may influence a number of intestinal ion transport processes that ultimately may influence water absorption: (1) reduced NaCl cotransport, (2) reduced Cl(-) uptake via Cl(-)/HCO(3(-)) exchange and (3) reduced precipitation of Ca(2+) and Mg(2+) carbonates. Despite these effects on intestinal ion and water transport, overall osmoregulatory status was not compromised in E2-treated fish, suggesting the possibility of compensation by other organs.

Brain cytochrome P450 aromatase activity in roach ( Rutilus rutilus): Seasonal variations and impact of environmental contaminants

P450 aromatase catalyses the conversion of C19 androgens to C18 estrogens which is thought to be essential for the regulation of the reproductive function. In this study, brain aromatase activity (AA) was measured monthly over a reproductive cycle in wild roach ( Rutilus rutilus) sampled in a reference site in Normandy. AA peaked during the breeding season, reaching 35 fmol mg −1 min −1 in both male and female fish, and was low during the rest of the year except for a significant rise in October. AA was correlated with ovary maturation (measured either as gonado-somatic index or by histological analysis of the gonads) and plasma sex-steroid levels (11-ketotestosterone in males and 17-β-estradiol in females). Measurements of AA in polluted sites showed that activity was significantly upregulated in sites with fish showing high levels of plasma vitellogenin and large proportion of intersexuality (20–50%) thus suggesting the occurrence of estrogenic compounds and their involvement in AA modulation.

Vitellogenin Induction and Reproductive Status in Wild Chilean Flounder Paralichthys adspersus (Steindachner, 1867) as Biomarkers of Endocrine Disruption Along the Marine Coast of the South Pacific

This study evaluated the condition factor, gonadosomatic, and hepatosomatic indexes, occurrence of plasmatic vitellogenin (Vg), and frequency of spermatogenic maturity stages in male Chilean flounders, Paralichthys adspersus, caught at three different coastal sites off the Bio-Bio region, central Chile, during 1 year. The Vg was detected by polyacrylamide gel electrophoresis with sodium dodecyl sulfate and Western blot analyses using an antibody against Chilean flounder Vg. The spermatogenic maturity stages were analyzed by histological gonadic diagnostic. The prevalence of plasmatic Vg induction in male fish differed significantly among sites. The flounders sampled from the Itata area were the most affected. Evaluations of biometric data, plasmatic Vg induction, and spermatogenic maturity stages of the flounder showed the following: (1) lower gonadosomatic index, (2) greater hepatosomatic index, (3) greater prevalence of plasmatic Vg, and (4) delayed development of the gonad. The results suggest that estrogenic endocrine-disruption compounds are introduced into the marine environment, negatively affecting the fish studied. The relevance of this report is discussed in relation to estrogenic compounds introduced by industrial and municipal wastewater effluents in the areas studied.

Life‐cycle exposure to the estrogenic mycotoxin zearalenone affects zebrafish (Danio rerio) development and reproduction

Zearalenone (ZON) is one of the worldwide most common mycotoxin and exhibits estrogenic activity in the range of natural steroid estrogens. The occurrence of ZON has been reported in soil, drainage water, wastewater effluents, and rivers, but its ecotoxicological effects on fish have hardly been investigated. The consequences of continuous long-term ZON exposure, including a subsequent depuration period, as well as transgenerational effects of F0 short-term exposure on F1 generation were investigated. Effects on growth, reproduction activity, physiology, and morphology of zebrafish (Danio rerio) were examined in a 182 day live-cycle experiment. Life-long exposure to ZON for 140 days increased wet weight, body length, and condition factor of female fish at 1000 ng/L, and sex ratio was shifted toward female from 320 ng/L ZON. Only females at 1000 ng/L ZON revealed a 1.5-fold induction of plasma vitellogenin (VTG). Relative fecundity at 1000 ng/L recovered significantly during the depuration period. An increased condition factor in adult female F1 fish implies that exposure of F0 generation to 1000 ng/L ZON affected growth of F1 generation. A negative correlation between relative fecundity in the F1 generation (all groups exposed to 320 ng/L ZON) and the nominal ZON concentrations of the F0 exposure might indicate an influence of F0 exposure on reproductive performance of F1 generation. No exposure scenario affected fertility, hatch, embryo survival, and gonad morphology of zebrafish. Evaluating the environmental relevance of this data, the risk for fish to be harmed by exposure to ZON solely seems rather marginal, but ZON might contribute to the overall estrogenicity in the environment. © 2011 Wiley Periodicals, Inc. Environ Toxicol 2013.

Development of an enzyme-linked diagnostic system for the detection of vitellogenin in the blood plasma of the barfin plaice Liopsetta pinnifasciata

Vitellogenin (Vg) of the barfin plaice Liopsetta pinnifasciata was isolated and purified. Using SDS-PAGE, barfin plaice Vg was dissociated into polypeptides with molecular masses of 180, 98, 70, 52, 41, and 37 kDa. Rabbit polyclonal antibodies to this protein were obtained and a specific highly sensitive diagnostic enzyme-linked immunosorbent assay was developed for Vg detection in the blood plasma of the barfin plaice, which was tested on fish from natural habitats. The plasma Vg concentration in L. pinnifasciata from a contaminated location of Amursky Bay (Sea of Japan) in November varied from 5.295 to 28.367 mg/ml in females and from an undetectable level to 0.957 mg/ml in males. The presence of Vg in the blood plasma of male L. pinnifasciata from Amursky Bay, which was detected for the first time, is indicative of ecosystem pollution by chemicals with estrogenic activity.

Alligator gar (Atractosteus spatula, Lacépède 1803) vitellogenin: purification, characterization and establishment of an enzyme-linked immunosorbent assay

Alligator gar (Atractosteus spatula) is a non-teleost bony fish distributed in North America. Gar populations have drastically declined as a consequence of habitat deterioration and the lack of regulation for their capture. Control of reproduction is critical for recovering their natural populations. The impossibility to distinguish genders and the determination of sexual maturity have hindered their successful reproduction. This research was aimed at developing an enzyme-linked immunosorbent assay (ELISA) for the quantitative estimation of vitellogenin (VTG), a female-specific protein. Plasmatic VTG from 17β-estradiol (E2)-induced juveniles and ovary vitellin from adult females were purified and characterized. Polyclonal antibodies against both proteins were produced to develop an ELISA. The immunoassay was validated by quality tests such as sensitivity, parallelism, recovery, reproducibility and specificity. Vitellogenin was determined in different tissues (plasma, mucus, liver and gills) of alligator gar. Vitellogenin and E2 concentrations in female breeders were found to be higher in November, before the spring spawning season. This approach represents a quick, reliable and non-invasive practical alternative to distinguish genders and evaluate gonad maturation.

Development of enzyme-linked immunosorbent assays for plasma vitellogenin in Chinese rare minnow ( Gobiocypris rarus)

Due to the wide occurrence of endocrine disrupting chemicals in the environment, it is much of importance to develop high throughput screening method for the analysis of this kind of pollutants. Using anion-exchange membrane chromatography, vitellogenin (VTG) from the plasma of 17β-estradiol (E 2) treated Chinese rare minnow was rapidly purified within 15 min. Both polyclonal antibody (PcAb) and monoclonal antibody (McAb) against rare minnow VTG (R-VTG) were prepared in rabbit and Balb/c mice, respectively. The competitive enzyme-linked immunosorbent assays (ELISA) based on either PcAb or McAb were developed to identify and quantify R-VTG in the plasma, and these two methods showed similar characteristics. The detection limits of both assays were lower than 3 ng mL −1 with the working ranges covering three magnitudes. The recovery efficiencies of PcAb and McAb based ELISA were 104.2% and 102.6%, respectively; and the intra-assay and inter-assay of these two assays were 6.2% and 9.2%, 8.6% and 12.8%, respectively. These results indicated that the described competitive ELISA methods were sensitive and valuable tools for quantifying vitellogenin in rare minnow plasma. These methods were then applied to measure R-VTG concentrations in plasma of male fish exposed to a series of E 2 concentrations. When E 2 levels were less than 10 ng L −1, R-VTG levels in plasma were comparable to that in solvent control, while R-VTG levels significantly increased 15-folds and 350-folds, respectively when E 2 exposure concentrations were controlled at 10 and 50 ng L −1. The high sensitivity of Chinese rare minnow to E 2 was demonstrated, making it a valuable model species to study environmental estrogens.

Enzyme-linked immunosorbent assay (ELISA) measurement of vitellogenin in plasma and liver histopathology in barfin plaice Liopsetta pinnifasciata from Amursky Bay, Sea of Japan

Vitellogenin (Vg) of the barfin plaice Liopsetta pinnifasciata was isolated and purified. In native polyacrylamide gel electrophoresis, Vg appeared as one band. After being subjected to sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE), Vg fraction produced several polypeptides with molecular masses of 180, 98, 70, 52, 41 and 37 kDa. MALDI-TOF mass spectrometry (MS) of the 180- and 98-kDa Vg polypeptides from the SDS-PAGE gel and de novo sequencing of their four peptide fragments based on MS/MS analysis confirmed that the purified proteins were vitellogenins, which shared high similarity with the Vgs of the barfin flounder Verasper moseri and Atlantic halibut Hippoglossus hippoglossus. The most part of the predicted sequences obtained from the L. pinnifasciata 180-kDa polypeptide has previously been found in the V. moseri vitellogenin type B, the sequences obtained from the 98-kDa polypeptide were found in V. moseri vitellogenin type A, so these findings allow us to propose that L. pinnifasciata has at least two different forms of Vg. Rabbit polyclonal antibodies against Vg were produced, and a quantitative enzyme-linked immunosorbent assay was developed. The concentration of Vg in barfin plaice from the moderately contaminated area of Amursky Bay in the Sea of Japan was detected based on the maturity stage of their gonads. In November 2008, the Vg concentration in the plasma of females with advanced oogenesis varied from 5.295 to 28.367 mg/ml (mean 16.38 ± 6.73 mg/ml, CV = 41.1%); in the plasma of males, the concentration ranged from non-detectable to 0.957 mg/ml (0.29 ± 0.42 mg/ml, CV = 127.9%). In October 2009, the Vg concentration in female plasma was lower than in November 2008 (2.21-13.87 mg/ml). High individual variability of plasma Vg was characteristic for maturing males (CV = 200.3%) and immature females (CV = 255.5%), and there was no significant difference between plasma Vg concentrations in males captured in November 2008 and October 2009 or in maturing males and immature females. Vacuolisation of hepatocytes was more typical for males with low plasma Vg concentrations and females with high plasma Vg concentrations. Necrosis and pyknosis of hepatocyte nuclei were more frequent in males with high Vg concentrations and in females with low plasma Vg concentrations.

Impact of photoperiod manipulation on day/night changes in melatonin, sex steroids and vitellogenin plasma levels and spawning rhythms in Senegal sole, Solea senegalensis

Photoperiod and temperature are known as the main synchronizers of seasonal reproduction in fish. This paper studied the role of photoperiod on the synchronization of F1 Senegal sole reproduction rhythms. Fish were maintained under constant short-photoperiod (9L:15D) from the winter solstice onwards (experimental group) or under naturally-changing photoperiod (control group), and water temperature naturally oscillated in both groups. Blood samples were collected during the reproduction season at pre-spawning (March), spawning (April) and post-spawning (May) to determine the endocrine status. Spawning events and egg quality parameters were also monitored. The results revealed a significant increase in nocturnal melatonin concentration from March to May in the control group, while in the experimental group such seasonal change did not occur. As to plasma levels of vitellogenin, testosterone, estradiol and 11keto-testosterone, differences between groups were found mostly in March, while in April and May levels were often similar. Spawning was observed in both groups, although the experimental group started slightly earlier and also finished earlier than the control group, perhaps as a result of the increase in sex steroids and VTG observed at pre-spawning. Briefly, reproduction rhythms persisted in the absence of the natural lengthening of photoperiod, although photoperiod manipulation altered the seasonal modulation of melatonin, increased sex steroids and vitellogenin at pre-spawning, and slightly advanced the timing of spawning.

Water quality evaluation of two interconnected dam lakes with field-captured and laboratory-acclimated fish, Cyprinus carpio

Karakaya and Sultansuyu Dam Lakes, located in the eastern part of Turkey, are important water sources, both for irrigation and fishery. The main goal of the study was to investigate water qualities of dam lakes using a set of biomarkers in the fish Cyprinus carpio. For this aim, field sample and laboratory-acclimated fish were compared to identify changes in selected biomarkers. The activities of ethoxyresorufin-O-deethylase, glutathione S-transferase, glutathione reductase, and carboxylesterase were determined in liver samples. Also, plasma and liver lactate dehydrogenase, aspartate aminotransferase, and alanine aminotransferase activities were assayed. Brain acetylcholinesterase and carboxylesterase activities were also determined. The hepatosomatic index and condition factors were calculated. Plasma vitellogenin assays were evaluated for the presence of xenoestrogen. Physicochemical values of water samples showed the existence of eutrophication risk, and also, some chemicals in both lakes were determined to be over tolerable limits. The comparisons of samples from both dam lake and laboratory-acclimated fish showed that the lakes may be at risk of pollution by some xenobiotics, namely xenoestrogens and acetylcholinesterase-inhibiting agents.