Sex Reversal In Fish Research Articles

Sexual plasticity is affected by sex chromosome karyotype and copy number of sex determiner in tilapia

Teleost fish exhibit remarkable plasticity of sexual differentiation as sex reversal can be induced by exogenous steroid hormones. To date, little is known about whether the sex chromosome karyotype and copy number of sex determiners affects sex reversal in fish. The Nile tilapia (XX/XY system with amhy as sex determiner) and blue tilapia (ZZ/ZW system) are two closely cichlids widely used for aquaculture. In this study, we measured the sex reversal rates of XY, YY-amhy+/+, YY-amhy+/−, ZX and ZZ tilapia by 17β-estradiol (E2) treatment, ZW and WW tilapia by aromatase inhibitor (letrozole) treatment and aimed to promote sex reversal of YY fish through blockage of androgen synthesis and administration of E2. Statistically, E2 treatment resulted in lower sex reversal rate in YY and ZZ fish compared with XY and ZX fish, respectively. In addition, letrozole treatment led to lower sex reversal rate in WW fish compared with ZW fish. Importantly, E2 treatment of YY fish with mutation of one copy of amhy resulted in similar sex reversal rate compared with XY fish. These results implied that the two homogametic sex chromosomes in YY and ZZ fish antagonized the E2 induced sex reversal due to the up-regulation of the male pathway genes, and in WW fish antagonized the effects of the letrozole induced sex reversal. Simultaneous treatment of E2 and trilostane, an inhibitor of 3β-hydroxysteroid dehydrogenase, promoted sex reversal of YY fish. Taken together, our results demonstrate that sexual plasticity is associated with the sex chromosome karyotype and copy number of the sex determiner in tilapia. The production of YY neofemale tilapia allowed establishment of a YY maintenance line and large scale breeding of XY genetically male tilapia in aquaculture.

Environmentally-induced sex reversal in fish with chromosomal vs. polygenic sex determination

Sex ratio depends on sex determination mechanisms and is a key demographic parameter determining population viability and resilience to natural and anthropogenic stressors. There is increasing evidence that the environment can alter sex ratio even in genetically sex-determined species (GSD), as elevated temperature can cause female-to-male sex reversal (neomales). Alarmingly, neomales are being discovered in natural populations of several fish, amphibian and reptile species worldwide. Understanding the basis of neomale development is important for conservation biology. Among GSD species, it is unknown whether those with chromosomal sex determination (CSD), the most common system, will better resist the influence of high temperature than those with polygenic sex determination (PSD). Here, we compared the effects of elevated temperature in two wild zebrafish strains, Nadia (NA) and Ekkwill (EKW), which have CSD with a ZZ/ZW system, against the AB laboratory strain, which has PSD. First, we uncovered novel sex genotypes and the results showed that, at control temperature, the masculinization rate roughly doubled with the addition of each Z chromosome, while some ZW and WW fish of the wild strains became neomales. Surprisingly, we found that at elevated temperatures WW fish were just as likely as ZW fish to become neomales and that all strains were equally susceptible to masculinization. These results demonstrate that the Z chromosome is not essential for male development and that the dose of W buffers masculinization at the control temperature but not at elevated temperature. Furthermore, at the elevated temperature the testes of neomales, but not of normal males, contained more spermatozoa than at the control temperature. Our results show in an unprecedented way that, in a global warming scenario, CSD species may not necessarily be better protected against the masculinizing effect of elevated temperature than PSD species, and reveal genotype-by-temperature interactions in male sex determination and spermatogenesis.

High temperature stress response is not sexually dimorphic at the whole-body level and is dependent on androgens to induce sex reversal

The understanding of the molecular and endocrine mechanisms behind environmentally-induced sex reversal in fish is of great importance in the context of predicting the potential effects of climate change, especially increasing temperature. Here, we demonstrate the global effects of high temperature on genome-wide transcription in medaka (Oryzias latipes) during early development. Interestingly, data analysis did not show sexual dimorphic changes, demonstrating that thermal stress is not dependent on genotypic sex. Additionally, our results revealed significant changes in several pathways under high temperature, such as stress response from brain, steroid biosynthesis, epigenetic mechanisms, and thyroid hormone biosynthesis, among others. These microarray data raised the question of what the exact molecular and hormonal mechanisms of action are for female-to-male sex reversal under high temperatures in fish. Complementary gene expression analysis revealed that androgen-related genes increase in females (XX) experiencing high water temperature. To test the involvement of androgens in thermal-induced sex reversal, an androgen antagonist was used to treat XX medaka under a high-temperature setup. Data clearly demonstrated failure of female-to-male sex reversal when androgen action is inhibited, corroborating the importance of androgens in environmentally-induced sex reversal.

Expression patterns of kiss2 and gpr54-2 in Monopterusalbus suggest these genes may play a role in sex reversal in fish.

Due to its exceptionally small genome size and protogynous hermaphroditism, Monopterus albus has been proposed as a model for vertebrate sexual development. The Kiss/GPR54 system is a central regulator of sexual development in most vertebrates, but its role in sex reversal remains hypothetical. In contrast to mammals, fishes often possess more than one copy of the kiss and gpr54 genes. Our objectives were to identify all kiss/gpr54 genes in the genome of M. albus and to assess their involvement in sex reversal via their expression patterns (qPCR) in females, males, and intersex specimens. We identified only two genes: kiss2 and gpr54‐2. kiss2 expression was extremely high in the gonads of males, intermediate in females, and low in intersex; and reduced in all tissues of intersex. gpr54 expression was also extremely high in the gonads of males, high in intersex, but low in females. gpr54 expression in brain was high in all three sexes. In conclusion, (a) kiss1 has been functionally replaced in M. albus; (b) the functions of gpr54‐2 in brain are not sex‐specific; (c) kiss2 appears to undergo a ‘reset’ in the expression during the sex change; and (d) sex‐specific expression patterns in the gonads indicate that these two genes may play a role in sex reversal in fish.

Production of Monosex All-Male Nile Tilapia Using Ethanol Extract of Tribulus terrestris Seeds

Mixed sex juveniles of Nile tilapia (mean weight 0.025 ± 0.009 g; mean length 1.25 ± 0.012 cm) were subjected to dietary treatment (commercial fish feed, Tokyu, Japan, 30% crude protein) with ethanolic extract of Tribulus terrestris seeds at different concentrations (0.0, 2.0, 2.5, 3.0 g/kg feed) to evaluate the efficacy of the plant seed extract for induction of masculinisation in fish. Treatment with plant extract at different concentrations have resulted in significantly higher (P < 0.05) survival percentage compared to control indicating no adverse health effect of the plant extract. Fish fed diets containing the plant extract showed significantly higher (P < 0.05) percentage of males compared to fish fed control diet. Dietary supplementation with the plant extract at the concentration of 2.0 g/kg feed showed the highest percentage (91.53 ± 0.38) of males and may be considered to be the best concentration among the selected concentrations for monosex tilapia production with T. terrestris seeds extract. The result of the study indicates that ethanolic extract of T. terrestris seeds may have potential for inducing sex reversal in fish and this biodegradable natural plant material may be used for monosex tilapia production instead of synthetic steroids.

The fitness consequences of environmental sex reversal in fish: a quantitative review

Environmental sex reversal (ESR) occurs when environmental factors overpower genetic sex-determining factors. The phenomenon of ESR is observed widely in teleost species, where it can be induced by exposing developing fish to endocrine disrupting chemicals (EDCs). EDC-induced ESR has been exploited by the aquaculture industry, while ecological and evolutionary models are also beginning to elucidate the potential roles that sex-reversed individuals play in influencing population dynamics. However, how EDC exposure affects individual fitness remains relatively unknown. To date, many experimental studies have induced sex reversal in fish and measured fitness-as indicated by related traits such as size, survival and gonadal somatic index (GSI), but the reported results vary. Here, we meta-analytically combine the results of 78 studies of induced ESR to gain insight into the fitness of sex-reversed individuals. Overall, our results suggest that the fitness of fish exposed to EDCs is reduced at the time of exposure, with exposed individuals having a smaller size and likely a smaller GSI. Given a period of non-exposure, fish treated with EDCs can regain a size equal to those not exposed, although GSI remains compromised. Interestingly, survival does not appear to be affected by EDC treatment. The published reports that comprise our dataset are, however, based on captive fish and the general small size resulting from exposure is likely to lead to reduced survival in the wild. Additionally, reduced fitness-related parameters are likely to be due to exposure to EDCs rather than ESR itself. We suggest that theoretical models of ESR should account for the fitness-related effects that we report. Whilst we are able to shed light on the physical fitness of EDC-exposed fish, the behaviour of such individuals remains largely untested and should be the focus of future experimental manipulation.

Molecular cloning and expression of TRα and TRβ in the protandrous cinnamon clownfish, Amphiprion melanopus during sex reversal

We divided the process of sex reversal from immature male to mature female in the protandrous cinnamon clownfish (Amphiprion melanopus) into six developmental stages as follows: I, immature male; II, mature male; III, male at 60 days after female removal; IV, male at 90 days after female removal; V, male at 120 days after female removal; and VI, mature female. Thyroid hormone receptors α (TRα) and β (TRβ) cDNAs were cloned from the ovary and mRNA expression levels were compared during the sex-reversal process. The nucleotide sequences of the TRα and TRβ cDNA were 1230 and 1188 bp in length with open reading frames encoding peptides of 409 and 395 amino acids, respectively. We observed that TRα mRNA and protein levels were high in all stages except the immature gonad, while TRβ mRNA levels were higher in the mature ovary than in any other gonadal stage. We injected gonadotropin-releasing hormone analogue to identify its effects on TRs mRNA in immature fish. The mRNA levels of TRs increased significantly. We therefore propose that TRs are related to testicular development as well as ovarian development in cinnamon clownfish. The present study also provides basic data on the role of TRs during sex reversal in fish.

A model for environmental sex reversal in fish

A mathematical model is presented which combines genetic XX-female/XY-male sex determination with environmental pressure for phenotypic sex reversal. This may occur when fishes are exposed to endocrine disrupters, specifically masculinization by exposure to androgens and feminization by exposure to estrogens. A generic model is derived for the sex ratio in successive generations and three special cases, with chronic and constant pressure to sex reverse, are discussed in detail. These show that, with extreme environmental pressure to masculinize, the male genotype is at risk of dying out but with less extreme pressure, masculinization will not be detectable since the proportion of phenotypic males becomes one-half. With feminization at any pressure to sex reverse, the male and female genotypes will be maintained in a stable sex ratio in which the proportion of genotypic males exceeds one-half and is close to one-half if YY offspring (eggs) are not viable. In converse, the model is also applicable to the genetic ZZ-male/ZW-female system of sex determination in fish. At present suitable data are not available with which to validate the model, but proposals are made for relevant experimental studies.

NOT ENOUGH MALES? MONOAMINES CHANGE EVERYTHING!

The saddleback wrasse, Thalassoma duperrey, inhabits corals reefs in the Hawaiian Islands. Remarkably, when the ratio of males to females in a population of wrasse becomes too low, the largest female transforms herself into a male over a 6–8-week period. Unsurprisingly, sex reversal in fishes requires a complete reorganization of physical, physiological and behavioural systems. In addition, sex reversal requires the conversion of external social cues into internal chemical cues. While past studies had investigated the roles of various hormones, neurotransmitters and neuromodulators as the trigger for the sex reversal process, an important group of chemicals that had not yet been investigated were the monoamine neurotransmitters such as dopamine, serotonin and noradrenaline. The hypothesis of the study by Larson and coworkers was that sex reversal in the wrasse would be accompanied by changes in the activities of these monoamines within different brain regions.In order to induce sex reversal in the larger fish and investigate the role of monoamines in the process, a series of floating enclosures were set up and a large and small female were placed in each. The team chose to monitor regions of the brain that are known to be involved in regulating sexual function and behaviour and are homologous to mammalian systems. The team sampled brains from the larger fish at various time points throughout the process of sex reversal and measured the monoamine levels using high-performance liquid chromatography (HPLC).Larson and colleagues found changes in monoamine metabolism in all of the brain regions that were examined. The most prevalent alterations in monoamine system activation were observed during the first week of sex reversal, at which time the sexual transition is primarily behavioural.One of the brain regions sampled appeared to be involved in regulating the fish's physical alterations. In the preoptic area of the brain, linked with male sexual function and behaviour in all vertebrates, the team found changes in both serotonergic activity and noradrenergic activity, which could potentially trigger the reorganization of the reproductive axis.The remaining four regions of the brain appeared to be involved in altering the female's behaviour during her sexual transition. The amygdala, an area in the brain that is important with respect to aggression, dominance and stress in vertebrates, exhibited significant changes in serotonin metabolism during sex reversal in wrasse. These changes were probably related to the observable increase in territorial acquisition and defense as the wrasse changed from female to male. In the ventral hypothalamus, noradrenergic activity, which is typically associated with female behaviour in this region of the brain, was reduced. By contrast, the locus coeruleus showed an increase in noradrenergic activity in the later stages of sex reversal, probably in response to an increase in circulating male sex hormones such as testosterone. The raphe nucleus showed an increase in serotonergic activity at the early, behavioural stages of sex reversal. During this time it is critical for the changing female to show dominance over other females, and dominant behaviour results in lower serotonergic activity in many animal models.The present study is the first to investigate monoamines and their role in environmentally stimulated sex reversal and has revealed that monoamines in different brain regions are important in both behavioural and gonadal sex reversal in the saddleback wrasse.

Recent Advances in Hormonal Induction of Sex-Reversal in Fish

ABSTRACT A neuroendocrine peptide can induce sex-reversal in pro-togynous labrid teleosts, similar to androgenic and antiestrogenic steroids. Culture water containing metabolites of 17a-methyltestosterone (MT), when recirculated, are more potent to induce sex-reversal than dietary supplemented MT; human estrogens present in sewage water, or other pollutants derail the normal course of sex differentiation. Steroid administration by immersion is 200-1,000 times cheaper than dietary supplementation; discrete immersion for 2-8 hours per day for 1-3 weeks after hatching induces maximal sex-reversal in salmonids like coho salmon, Oncorhynchus kisutch, while others like chinook salmon, O. tshawytscha, require dietary supplementation. Studies on uptake, disposition in the organs, and elimination of unlabeled or labeled steroids have indicated that the androgen is absorbed within one hour after intake, and selectively retained it in gall bladder, pyloric caeca and liver at higher concentrations for a longer time; however, 90-95% of the steroid is eliminated within 1-2 weeks. Estimated residual steroid of < 5 ng/g fish is too low to be a hazard to human health. Steroid delivered through water can be absorbed and retained by egg yolk of salmonids and is gradually released into the developing embryos. Temperature and autosomal gene(s) alter the process of sex differentiation in hormonally sex-reversing fish. Suboptimal treatment of steroids results in the production of hermaphroditic and intersex individuals. Hormonally sex-reversed fish also suffers from low survival and functional deficiences. The Indian scenario on the use of hormones for monosex aquaculture is described.

Xenoendocrine disrupters: laboratory studies on male reproductive effects

Wildlife from ecosystems contaminated with endocrine disrupting chemicals (EDCs) display a variety of reproductive alterations including cryptorchidism in the Florida panther, small baculum in young male otters, small penises in alligators, sex reversal in fish, and altered social behavior in birds. It has been proposed that EDCs also may have contributed to increases in testicular cancer and hypospadias and the reported decline in human sperm counts. Combined in vivo and in vitro studies are necessary to fully characterize EDC induced alterations of reproduction. We have identified several pesticides (vinclozolin, procymidone, p,p′-DDE) which bind rat and human androgen receptors, block androgen-induced gene expression in vitro and in vivo, delay puberty, reduce sex accessory gland size and alter sex differentiation in the male rat. Some of the phthalates, which are estrogenic in vitro but not in vivo, cause malformations in male rats that appear to result from antagonism of androgens in utero. In contrast, xenoestrogens affect male offspring but they are not malformed or infertile. Prenatal administration of an Ah receptor agonist (2,3,7,8-TCDD or PCB 169) produces a different spectrum of effects including reduced ejaculated sperm numbers in male rats.

The value of mechanistic studies in laboratory animals for the prediction of reproductive effects in wildlife: Endocrine effects on mammalian sexual differentiation

AbstractWildlife populations from contaminated ecosystems display a variety of reproductive alterations, including cryptorchidism in the Florida panther, small baculum in young male otters, small penises in alligators, sex reversal in fish, and altered social behavior in birds. The formation of biologically plausible hypotheses regarding disruption of reproduction in wildlife can be facilitated by mechanistic studies on laboratory animals. To this end, we are investigating the in vivo and in vitro effects of endocrine‐disrupting toxicants in rodents. In vitro studies have used receptor binding and transfected cell assays to confirm the suspected mechanism of action, whereas in vivo rodent studies examine altered sexual differentiation. Antiandrogenic pesticides compete with the natural ligands for both rat and human androgen receptors, block androgen‐induced gene expression in vitro and in vivo, delay puberty, reduce sex accessory gland size, and alter male rat sex differentiation. In contrast, xenoestrogens affect female central nervous system sex differentiation and fecundity without producing malformations or infertility in male offspring. Prenatal administration of 2,3,7,8‐tetrachlorodibenzo‐p‐dioxin (TCDD) or the TCDD‐like polychlorinated biphenyls produce yet another profile of effects in the offspring, reducing numbers of ejaculated sperm in male progeny and inducing urogenital malformations in females. Although phthalates are reported to be estrogenic in vitro, in vivo exposure causes developmental alterations that more closely resemble antiandrogenic activity. The mammalian data indicate that exposure to endocrine‐disrupting chemicals produces effects that are pathognomonic for mechanisms by which they act. Mechanistic information derived from mammalian studies can enhance our ability to predict toxicant effects on reproduction in fish and wildlife.