Intersex Fish Research Articles

Reproductive health of brown trout inhabiting Swiss rivers with declining fish catch

In recent years, brown trout catches have been declining in many Swiss rivers. One hypothesis is that this declining catch is linked to environmental estrogens, known to have the potential to adversely affect fish reproduction. In order to assess if the reproductive health of brown trout is impaired, we sampled fish at three sites along four rivers with a well documented catch decline. These rivers are affected by inputs of treated sewage effluent. The sampling was conducted during two years; we measured plasma vitellogenin (Vtg) concentrations and surveyed gonadal histology. Analysis of plasma Vtg revealed elevated concentrations (up to 8 µg/mL) in only 10 out of 197 males. Furthermore, there were no site specific patterns to this induction. These results indicate that the exposure to estrogenic compounds is low. Also the incidence of ovarian atresia was low and we found no male intersex fish. In contrast to males, females caught along two rivers had spermatogenic activity in ovarian tissue. However, this intersex condition does not appear to be connected to exposure to environmental estrogens. At one of 12 sites there was a high incidence of gonadal parasites in ovarian tissue, which may affect reproductive output but was not a general problem across sites. In conclusion, the exposure to estrogenic compounds does not appear to significantly affect the reproductive parameters we investigated in Swiss brown trout.

Masculinization of African Catfish (Clarias gariepinus) Treated with Gokshura (Tribulus terrestris)

The present paper describes the effects of Tribulus terrestris, a masculinization agent, on the sex reversal of African catfish, Clarias gariepinus. One-day-old hatchlings were immersed in water containing an extract of T. terrestris at a rate of 0, 3, 6, or 9 g/30 l water for 30 days. The 9 g treatment resulted in 80.42% male populations. Morphological and histological examinations of the gonads in all groups revealed no intersex fish. Histological examination of fish treated with T. terrestris revealed no damage to the testes or ovaries. Despite differences in sex ratio, growth rate, ovaries, and testes, fish treated with T. terrestris were histologically similar to fish from the control groups. Survival ranged from 72% in the 3 g group to 80% in the 9 g during treatment and did not significantly differ from the control (p>0.05). The best growth was obtained in the 9 g treatment.

Studies of Uptake, Elimination, and Late Effects in Atlantic Salmon (Salmo salar) Dietary Exposed to Di-2-Ethylhexyl Phthalate (DEHP) During Early Life

The phthalate esters are a group of industrial chemicals considered to have endocrine-disrupting properties. The most common tonnage product among these, di-2-ethylhexyl phthalate (DEHP), is widely spread in the environment. The objectives with the present work were to study uptake and metabolism of orally administered DEHP and its major metabolite mono-2-ethyl hexyl phthalate (MEHP) and to evaluate the impact of early life exposure on sex differentiation in Atlantic salmon. The feeding with contaminated diet started immediately after yolk sac resorption and continued for 4 weeks. Nominal concentrations of DEHP in the diet were 400 (measured 359), 800 (measured 827), and 1500 (measured 1648) mg DEHP/kg and a control group was fed food mixed with solvent. After the exposure period, fish were fed non-contaminated diet until final sampling 4 months post-exposure. There were no effects on growth or survival of the fish and no late effects on hepatosomatic index or sex ratio. However, the histological examination of gonads from fish exposed to 1500 mg DEHP/kg revealed a small but significant incidence (3%) of intersex fish (ovo-testis). Chemical residues of DEHP and MEHP were analyzed weekly during the first 3 months of the post-exposure period. Both DEHP and MEHP were rapidly eliminated to near background levels within one week post exposure. The study indicates that exposure of Atlantic salmon to relatively high concentrations of DEHP during a sensitive part of the life cycle may interfere with gonad differentiation.

Male-Biased Sex Ratios and Vitellogenin Induction in Zebrafish Exposed to Effluent Water from a Swedish Pulp Mill

Juvenile zebrafish (Danio rerio) were exposed to different dilutions (0, 0.67, 2.5, 10, and 50%) of effluent water from a Swedish pulp mill that previously has been reported to be androgenic to fish. Exposure was performed between days 10-38 days post-hatch. Fish were sampled for whole-body vitellogenin concentrations at day 38 post-hatch and for histological examination of gonads at day 60 post-hatch. In fish exposed to the highest concentration of pulp mill effluent, elevated concentrations of vitellogenin were measured. The androgenicity of the pulp mill water was confirmed by the increased number of males recorded at 60 days post-hatch. Image analysis of testes indicated stimulation of spennato genesis. Intersex fish were observed in all exposure groups. An androgenic activity equivalent to 5.6 ng/L dihydroxytestosterone was measured using the yeast androgen screen (YAS) assay. The present study demonstrates that both androgenic and estrogenic effects can be detected when exposing zebrafish during the juvenile period to complex mixtures of chemicals.

Prelude to Intersex in Fish: Identifying a Sensitive Period for Feminization

Field studies have shown a high occurrence of intersex (the presence of both male and female characteristics) and ovotestis (the presence of eggs in the testis) in wild populations of a fish known as roach (Rutilus rutilus) in rivers in the United Kingdom that are downstream from wastewater treatment plants. Furthermore, studies have demonstrated that intersex males are less fertile, which may have population-level effects. However, to date, scientists have been unable to induce intersex in male fish with controlled exposures to wastewater effluents. A study conducted at The University of Exeter now shows that the sensitive period for feminization of the reproductive duct—in which the male testis forms an ovary-like cavity—may occur earlier than previously thought, and raises new questions about the conditions that lead to actual germ cell disruption [EHP 113:1299–1307]. Many questions persist about the causes of and the most vulnerable life stages for various types of sexual effects induced by estrogenic chemicals in wastewater effluent. In this study, the researchers collected two different U.K. wastewater effluents and exposed wild roach at two life stages: during early life and development of the gonads (from fertilization up to 300 days post-hatch) and as adults producing germ cells following annual spawning. These adults included one group of fish that had been raised in clean water and another that had hatched and grown to maturity in the wild. Both effluents induced synthesis of vitellogenin (an estrogen-dependent yolk precursor and biomarker of estrogen exposure) at both life stages, with the extent of this induction correlating with the steroid estrogen content of the effluent. Previous studies have demonstrated that feminization of the sperm duct to form an ovary-like cavity occurs when exposure to effluent comes during the time of sexual differentiation, which in roach occurs from 50 to 150 days post-hatch. This study showed alteration of the sperm duct with an exposure earlier in life, from fertilization to 60 days post-hatch, before any signs of sexual development appear. The alteration, furthermore, was permanent, persisting even after 240 days’ maintenance in clean water after exposure. However, no ovotestis was observed in any of the juvenile fish. There was also no evidence of ovotestis in post-spawning adult male roach raised in a clean environment and subsequently exposed to effluent. There was evidence that the wild males had previously been exposed to estrogenic stimuli, as some of males had ovotestis when the study began. The severity of this condition increased slightly during the study period, but the increase occurred across both exposed and control fish and thus appeared unrelated to the study effluent exposure. The authors suggest possible explanations that need further study—one is that ovotestis is induced only by effluents with greater levels of estrogenic chemicals than those used in the study. The researchers evaluated the effluents for content of two chemicals previously implicated in causing intersex—steroidal estrogens and alkylphenols—and found that these levels were similar to concentrations reported in wastewater effluents in the United Kingdom and worldwide. They emphasize that chemical content and interactions ideally should be taken into account when trying to determine the conditions that lead to sexual effects. The results of these studies raise the possibility that ovotestis may be a result either of longevity of exposure or of programming in early life that manifests itself as fish mature sexually. Previous findings from the authors support this idea by showing that the severity of intersex increases with age. The authors are further exploring these possibilities now with a laboratory study of roach that includes an environmentally relevant estrogen exposure of two years’ duration.

A ranking system for the evaluation of intersex condition in european flounder (Platichthys flesus)

Intersex (ovotestis) is the condition whereby an individual possesses oocytes, at varying degrees of severity, within the normal testicular tissue and is regarded as an endpoint of endocrine disruption in male fish. The occurrence of intersex has been linked with exposure to endocrine-disrupting compounds present in the environment, which are thought to mimic the actions of the female sex hormone 17-beta-oestradiol. Using stained histological sections of archived cases of intersex from European flounder (Platichthys flesus), this study developed a grading method to assess the extent of the condition. This was termed the ovotestis severity index (OSI). The OSI utilizes the distribution pattern (focal, diffuse, cluster, zonal) and type of oocyte (previtellogenic, cortical alveolar, vitellogenic) to assign a severity rating to each intersex fish. A high OSI refers to fish with testicular material almost completely replaced with mature (vitellogenic) oocytes, and lower scores indicate those fish with small numbers of immature (previtellogenic) oocytes. The development of this index provides pathologists with a robust tool for the grading of the intersex condition in European flounder and potentially in other fish species.

Histological evidence of intersex in feral sharptooth catfish (Clarias gariepinus) from an estrogen‐polluted water source in Gauteng, South Africa

This is the first histological evidence of intersex in a fish species inhabiting a South African water source. One hundred catfish (Clarias gariepinus) were collected randomly from the Marais Dam (MD) and the Rietvlei Dam (RVD) in the Rietvlei Nature Reserve (RNR), South Africa. These dams drain water from a stream that receives effluent from industrial sites, agricultural activities, informal settlements, and municipal treatment plants. Each fish was evaluated macroscopically and had blood drawn, and its gonads were macroscopically and histologically examined to verify intersex potentially related to endocrine disruption. Gonadal histology of several fish showed primary oocytes scattered in testicular tissue, indicative of intersex. The results showed intersexuality in 20% of fish from both the dams. The GSI value for intersex fish was closer to male GSI values, suggesting that the sampled intersex fish were more likely to have occurred from the feminization of male catfish. Target chemical analyses showed that the water, sediment, and serum samples tested positive for p-nonylphenol (p-NP). The p-NP level in water and sediment at MD was 6360 and 4.0 microg/kg, respectively, whereas in sediment at RVD it was 113 microg/kg. Commonly found in the effluent from sewage treatment plants, p-NP in water and sediment indicates estrogenic water pollution, which might affect wildlife and humans dependent on these sources.

Sediments are major sinks of steroidal estrogens in two United Kingdom rivers.

The occurrence of intersex fish in a number of European rivers has been attributed to exposure to estrogenic chemicals present in sewage treatment work (STW) effluents. To further understand the environmental fate of these contaminants, the estrogenic activity of effluents, water, and sediments were investigated both upstream and downstream of the major STW discharge in two United Kingdom rivers. Estrogenic activity, determined using the yeast estrogen-receptor transcription screen, of the major STW effluents on both rivers was similar, ranging from 1.4 to 2.9 ng 17beta-estradiol equivalents (EEQ)/L. Estrogenic activities of surface waters 1 km upstream and downstream of both STW inputs were less than the limits of detection (0.04 ng/L); however, levels of estrogenic activity in sediments were between 21.3 and 29.9 ng EEQ/kg and were similar at both upstream and downstream sites. Effluent and sediment extracts were fractionated by reverse phase-high-performance liquid chromatography, and estrogenic active fractions were further analyzed by gas chromatography-mass spectrometry. The major active chemicals in the two effluents and in the sediments were estrone with lesser amounts of 17beta-estradiol; however, at one site, a number of other unidentified estrogenic fractions were detected in the sediments. These results suggest that riverine sediments are a major sink and a potential source of persistent estrogenic contaminants.

Production of monosex male black crappie, Pomoxis nigromaculatus, populations by multiple androgen immersion

Monosex populations can solve the problem of stunted black crappie populations in small impoundments and can increase the potential of black crappie as an aquaculture species. In this study, we investigated the effective mode and duration of exogenous androgen administration to produce monosex male black crappie populations. We conducted two experiments using the synthetic androgens, 17α-methyltestosterone (MT) and trenbolone acetate (TBA). In the first experiment, the same age (45 days old), two different size fry with mean total lengths (±S.D., N=50) of 20.1±1.4 and 26.1±2.0 mm were either fed 60 mg MT/kg diet for 45 days or immersed in a 1 mg MT/l solution on 10 occasions for 5 h/day every 3–5 days between 45 and 86 days post-hatch (dph). In the second experiment, a different cohort of fry was immersed in a 1 mg TBA/l solution for 5 h/day every 3–5 days either on seven occasions between 40 and 66 dph or on 10 occasions between 45 and 86 dph. Mean total lengths (±S.D., N=50) of fry at 40 and 45 dph were 20.3±0.9 and 21.6±1.2 mm, respectively. Both modes of MT administration were ineffective for altering sex ratios in larger black crappie fry. Oral administration of MT to smaller fry resulted in 23% intersex fish. MT administration via short periodic baths of smaller fry produced 96% male populations. TBA was as potent as MT to induce masculinization in black crappie: both immersion treatments produced all male populations. Results of the present study indicated that initial size of fry as well as initial age should be considered when selecting appropriate size black crappie fry for hormone treatments. Monosex male populations in black crappie can effectively be produced through a series of (seven or less) short (5 h) immersions of young fry (40−45 days old and 20−21 mm) in a 1 mg/l aqueous solution of MT or TBA. Besides high effectiveness, immersion method will be the preferred mode of androgen administration to black crappie because it reduces variation in hormone uptake associated with the oral administration of steroids and provides more flexibility in feeding during the period of gonadal differentiation.

Hormonal effects on sex differentiation and growth in sunshine bass Morone chrysops× Morone saxatilis

During the production of sunshine bass, the fish are graded several times to avoid loss due to cannibalism. Such grading of fish that may exhibit sexually dimorphic growth could result in changing the sex ratio of populations raised for food production. A preferred gender exists for most aquaculture species. We have found that, during the first year, male sunshine bass appear to grow faster than females; however, females grow faster during the second year and are usually heavier when the fish are harvested. Feeding 17-α-methyltestosterone did not increase the growth of fingerling fish; however, feeding estradiol decreased growth, apparently by decreasing feed consumption. Immersion treatment of 4-day-old fry for up to 8 h with 17α-methyltestosterone or estradiol did not change the sex ratio. Treatment of 60- or 103-day-old fish for 40 days with estradiol in the feed resulted in all female fish. 17α-methyltestosterone treatment during the earlier period resulted in intersex fish, and treatment of 103-day-old fish did not change the sex ratio. Direct feminization with estradiol can be achieved during a period of development in which direct masculinization with 17α-methyltestosterone is not effective.

Gonad development and vitellogenin production in zebrafish ( Danio rerio) exposed to ethinylestradiol and methyltestosterone

In a partial life-cycle test, the impact of 17α-ethinylestradiol (EE2) and 17α-methyltestosterone (MT) on juvenile zebrafish was evaluated by use of vitellogenin measurements and gonadal development. Exposure to EE2 (1–25 ng/l) resulted in a dose-dependent increase in vitellogenin production starting at 2 ng/l. Significant changes in sex ratios in female direction were detected at 1 ng/l, with complete sex reversal taking place after exposure to 2 ng/l. No intersex fish were observed after exposure to EE2. Exposure to MT resulted in decreased vitellogenin concentrations. Complete sex reversal was detected in all MT concentrations used (26–1000 ng/l). A large proportion of intersex fish was observed after exposure to 1000 ng MT/l. The period of gonadal sex reversal in non-exposed zebrafish was also studied. The main morphological features of the transformation of ovaries into testis were observed 4–5 weeks after hatching.

Successful Sex Reversal of Brook Trout with 17α‐Methyldihydrotestosterone Treatments

AbstractMonosex female stocks are widely used in the commercial production of rainbow trout Oncorhynchus mykiss. The potential for commercial production of brook trout Salvelinus fontinalis, however, is constrained by the lack of published protocols for producing the sex‐reversed males required to create monosex female stocks. Immersion and immersion plus feeding treatments with 17α‐methyltestosterone (MT) and 17α‐methyldihydrotestosterone (MDHT) were applied to genotypically female gynogenetic brook trout to induce phenotypic sex reversal. The fry were exposed to a 6‐h immersion in a solution of MT or MDHT on day 10 following completion of hatch and/or to a steroid‐treated diet for 60 d beginning at first feeding. Immersion dosages were 0.5 or 1.0 mg/L, and feeding dosages were 1.0 or 2.0 mg/kg of feed for MT and 0.5 or 1.0 mg/kg for MDHT. Phenotypic sex of the fish was determined 19 or 22 months after first feeding. Control gynogenetic fish were 100% phenotypic females. Treatments with MT had minimal effect: most fish remained female, with only a low incidence of phenotypic males (1–3% in four of the treatments), intersex fish, or sterile fish. In contrast, a substantial number of phenotypic males were observed in several of the MDHT treatments, with the highest proportion (45%) occurring in the 0.5 mg/L immersion plus 0.5 mg/kg feeding treatment. Sperm was obtained from 29 males from five MDHT treatment groups and one MT treatment group examined at maturity (22 months) and was used in progeny tests of these males. The progeny were 100% female, confirming the male parents to be genotypically female. These protocols may be used to create sex‐reversed brook trout males for the production of monosex female progeny, although additional trials are ongoing to test similar MDHT immersion dosages applied once or multiple times, with or without feeding treatments, to identify protocols with increased efficacy.

Wild intersex roach (Rutilus rutilus) have reduced fertility.

Endocrine-disrupting chemicals, known to be present in the environment, have great potential for interfering with reproductive health in wildlife and humans. There is, however, little direct evidence that endocrine disruption has adversely affected fertility in any organism. In freshwater and estuarine fish species, for example, although a widespread incidence of intersex has been reported, it is not yet known if intersexuality influences reproductive success. The purpose of this study was, therefore, to determine gamete quality in wild intersex roach (Rutilus rutilus) by assessing sperm characteristics, fertilization success, and ability to produce viable offspring. The results clearly demonstrate that gamete production is reduced in intersex roach. A significantly lower proportion of moderately or severely feminized fish (17.4% and 33.3%, respectively) were able to release milt compared with normal male fish from contaminated rivers (in which 97.6% of the males were able to release milt), reference male fish (97.7%), or less severely feminized intersex fish (experiment 1: 85.8%, experiment 2: 97%). Intersex fish that did produce milt produced up to 50% less (in terms of volume per gram of testis weight) than did histologically normal male fish. Moreover, sperm motility (percentage of motile sperm and curvilinear velocity) and the ability of sperm to successfully fertilize eggs and produce viable offspring were all reduced in intersex fish compared with normal male fish. Male gamete quality (assessed using sperm motility, sperm density, and fertilization success) was negatively correlated with the degree of feminization in intersex fish (r = -0.603; P < 0.001) and was markedly reduced in severely feminized intersex fish by as much as 50% in terms of motility and 75% in terms of fertilization success when compared with either less severely feminized intersex fish or unaffected male fish. This is the first evidence documenting a relationship between the morphological effects (e.g., intersex) of endocrine disruption and the reproductive capabilities of any wild vertebrate. The results suggest that mixtures of endocrine-disrupting substances discharged into the aquatic environment could pose a threat to male reproductive health.

Seasonal variation of plasmatic and hepatic vitellogenin and EROD activity in carp, Cyprinus carpio, in relation to sewage treatment plants

Concern about the health of aquatic fauna living in waters containing biologically active levels of estrogenic compounds is particularly focused on the effects on their reproductive success. To that end, carp, Cyprinus carpio, a feral fish living in warm waters of Southern Europe (NE Spain), were selected for signs of estrogenicity. The study area covered two tributaries (the Anoia and the Cardener) of the Llobregat River both known to be polluted by estrogenic compounds. The estrogenicity in the carp was measured as vitellogenin (VTG) presence in males and alterations in VTG levels in females, over a 6-month period, embracing both the pre- and post-spawning seasons. VTG content was measured in both the plasma and liver, the latter being the organ that synthesizes it. Also, hepatic 7-ethoxyresorufin O-deethylase (EROD) activity was recorded, as interactions of xenoestrogens and oestradiol have been reported to affect this enzymatic activity. The estrogenicity of these rivers was more evident in the Anoia at the location downstream from the sewage treatment plant (STP), by elevated levels of VTG in males and by the presence of some intersex individuals. In the Cardener, no intersex fish were found and male plasmatic VTG was not so highly elevated. However presence of hepatic VTG, in up to 54% of the male fish analyzed, proved exposure to xenoestrogens. In females, VTG fluctuated according to the biological cycle with a plasmatic peak in May and an earlier maximal in the liver. However, this pattern was altered in the locations with higher xenoestrogens presence. EROD activity showed differences between sexes, with higher activity in males than females, as well as site-related differences (up to one order of magnitude) in the same river. These differences were even greater than those detected between rivers. A seasonal trend was also seen in EROD activity with higher induction towards the summer in both males and females.

Altered sexual maturation and gamete production in wild roach (Rutilus rutilus) living in rivers that receive treated sewage effluents.

Disruption in gonadal development of wild roach living in U.K. rivers receiving large volumes of treated sewage effluent is manifest in a variety of ways, ranging from malformation of the germ cells and/or reproductive ducts to altered gamete production. Intersex fish were also found to have an altered endocrine status and an elevated concentration of plasma vitellogenin. Gonadal growth was inhibited only in severely intersex fish, whereas progression of spermatogenesis was delayed in a large proportion of all intersex and exposed male fish. In contrast to the effects observed in the intersex and exposed male fish, the maturation of ovaries in female fish inhabiting effluent-contaminated rivers appeared to be less obviously affected, although a higher incidence of oocyte atresia was found in the effluent-exposed fish compared with the reference fish. A positive correlation was found between the proportion of female tissue in the gonads of intersex fish and their plasma vitellogenin concentration, suggesting that vitellogenin can be an indicator for the level of gonadal disruption in intersex roach. The estradiol-17beta concentration in intersex fish was intermediate between the concentration found in males and females, and the plasma testosterone was between 2- and 3-fold higher in intersex fish compared with male fish. These data suggest a link between altered endocrine status in intersex and female fish and gonadal disruption. Spermiation was also affected in roach living in effluent-impacted rivers: a lower proportion of fish were found releasing sperm, and in those intersex fish that were spermiating, a reduced milt volume and a reduced sperm density were found. All intersex fish had malformations of the reproductive duct(s), and in severely affected fish, the ducts were occluded, thus preventing release of gametes. In view of the widespread occurrence of intersexuality in wild fish populations in rivers throughout the United Kingdom, assessment of the reproductive capabilities of these intersex roach is clearly needed to understand the impact of this phenomenon on roach fertility.

Paradoxes in exogenous androgen treatments of bluegill

Three experiments were conducted to investigate the effect of oral administration of trenbolone acetate and 17a-methyltestosterone on sex determination in 28-day-old bluegill, Lepomis macrochirus Rafinesque, fry. Multiple androgen doses and various treatment durations were tested. All treatments produced fewer males and females than did the control group (P < 0.01). All concentrations of both androgens produced a high proportion of intersex fish (38–81%). The number of males and females declined with the increase in androgen dose or treatment duration. Sterile fish were found in treatments with a higher dose rate or a longer treatment period. The predominance of intersex fish and reductions of both males and females in the androgen treatments suggest that gonadal development of both genotypic male and female fish were being altered by the hormone treatment.

Hormonal Sex Reversal and Evidence of Female Homogamety in Black Crappie

Abstract This study reports the results on hormone-induced sex reversal in black crappie Pomoxis nigromaculatus and the sex ratios of progenies obtained from test crosses of androgen-treated males with normal females. The androgen 17α-methyltestosterone (MT) was orally administered to fish with an artificial diet (30 mg/kg) for 30 d beginning 37 d after hatching; mean fish lengths were 35 and 55 mm at the beginning and conclusion of the treatment, respectively. The MT-treated group consisted of 95% males and 5% intersex fish, while the sex ratio in the control group was not significantly different from 1:1. Four males from the androgen-treated group were individually test-crossed with normal females to identify sex-reversed males according to the sex ratios in progenies and to reveal the chromosomal mechanism of sex determination in black crappie. Two out of three analyzed progenies consisted of females only, while one progeny had a sex ratio not significantly different from 1:1. The appearance of all-fem...

Sexual disruption in a second species of wild cyprinid fish (the gudgeon,Gobio gobio) in United Kingdom Freshwaters

To establish whether the intersex condition seen in the roach (Rutilus rutilus) in United Kingdom (UK) rivers was species specific or a more general phenomenon in fish, evidence for sexual disruption was sought in a second cyprinid species, the gudgeon (Gobio gobio). Gudgeon were collected from the Rivers Aire and Lea (at locations that receive high-volume discharges of sewage treatment works [STW] effluent and that contain intersex roach) and from two still waters, and their gonads were examined histologically for evidence of intersexuality (the simultaneous presence of oocytes and testicular tissue). Intersex gonads were found at all sites, with the highest incidences occurring at one of the still waters (Lakeside Fisheries: 15%) and at sites on the River Aire (Thwaite Weir, Silsden Bridge, and Knostrop: 14, 13, and 12%, respectively). In the River Lea and Longton Park Lake, the incidence of intersexuality in gudgeon was 6%. In most cases, intersex gonads were characterized by a few primary oocytes/gonad section in an otherwise normal testis. However, at some sites on the River Aire (Thwaite Weir and Knostrop), the intersex condition was more severe. At Thwaite Weir, for example, more than half of the gonad in 40% of the intersex fish was comprised of ovarian tissue. Elevated concentrations of plasma vitellogenin both in male and in intersex fish indicated that fish had been exposed to estrogen(s). Some of the gudgeon were found at sites several kilometers downstream of any point discharge of STW effluent; therefore, the results likely are representative of this species in wild populations found in typical UK river ecosystems. Together with the findings in the roach, these data on the gudgeon confirm that sexual disruption in fish in UK rivers is not species specific.

Sex reversal in Nile tilapia (Oreochromis niloticus) using a nonsteroidal aromatase inhibitor.

Several studies have demonstrated that steroid hormones can influence sex differentiation in nonmammalian vertebrates and it has been hypothesized that male and female sex differentiation are driven by androgen and estrogen hormones, respectively. Estrogen biosynthesis is mediated by the steroidogenic enzyme cytochrome P450 aromatase, which converts androgens to estrogens. In the present study we examined the efficacy of a potent nonsteroidal aromatase inhibitor incorporated into the food, on sex reversal of Nile tilapia (Oreochromis niloticus) larvae. Nile tilapia larvae were divided in seven groups, which were fed with diets containing different amounts of the aromatase inhibitor Fadrozole (0, 50, 75 and 100 mg/kg) during 15 and 30 days, starting 9 days after hatching. Independent of the period, the proportion of males was significantly higher in the treated groups. Treatment with the highest doses (75 and 100 mg/kg) for 30 days produced 100% males. Histological examination revealed no differences in gonadal tissues between control males and treated fish. Furthermore, one intersex fish was identified in the group treated with 50 mg Fadrozole/kg for 30 days. This study reports that a 100% Nile tilapia male population can be obtained by suppressing aromatase activity and suggests that besides steroid hormones, nonsteroidal compounds, such as aromatase inhibitors, have potential for production of monosex population in tilapia. J. Exp. Zool. 290:177-181, 2001.

A histological description of intersexuality in the roach

This paper is an illustrative guide to intersex in the roach Rutilus rutilus, based on 150 intersex individuals. Most intersex roach had female germ cells, or oocytes, within a predominantly male gonad (testis), and/or malformed/intersex reproductive ducts. The number, pattern and developmental stage of oocytes within testicular tissue in intersex fish varied greatly. In most intersex fish, a few primary oocytes, or numerous primary and secondary oocytes, were scattered randomly throughout the testicular tissue (multifocal intersex). In other, more severely feminized individuals, large areas of ovarian tissue were separated clearly from testicular tissue (focal intersex). Almost all intersex individuals had a female‐like reproductive duct (ovarian cavity). In mild cases of intersex (in which the majority of the germ cells were male) the ovarian cavity was present together with the male sperm duct/vas deferens, whilst in certain severe cases, the sperm duct was absent or vestigial.