Process Of Sex Reversal Research Articles

Protein expression patterns and metal metabolites in a protogynous hermaphrodite fish, the ricefield eel (Monopterus albus)

BackgroundThe ricefield eel Monopterus albus undergoes a natural sex change from female to male during its life cycle, and previous studies have shown the potential mechanisms of this transition at the transcriptional and protein levels. However, the changes in protein levels have not been fully explored, especially in the intersexual stage.ResultsIn the present study, the protein expression patterns in the gonadal tissues from five different periods, the ovary (OV), early intersexual stage gonad (IE), middle intersexual stage gonad (IM), late intersexual stage gonad (IL), and testis (TE), were determined by untargeted proteomics sequencing. A total of 5125 proteins and 394 differentially expressed proteins (DEPs) were detected in the gonadal tissues. Of the 394 DEPs, there were 136 between the OV and IE groups, 20 between the IM and IE groups, 179 between the IL and IM groups, and 59 between the TE and IL groups. Three candidate proteins, insulin-like growth factor 2 mRNA-binding protein 3 isoform X1 (Igf2bp3), triosephosphate isomerase (Tpi), and Cu-Zn superoxide dismutase isoform X1 [(Cu-Zn) Sod1], were validated by western blotting to verify the reliability of the data. Furthermore, metal metabolite-related proteins were enriched in the IL vs. IM groups and TE vs. IL groups, which had close relationships with sex change, including Cu2+-, Ca2+-, Zn2+- and Fe2+/Fe3+-related proteins. Analysis of the combined transcriptome data revealed consistent protein/mRNA expression trends for two metal metabolite-related proteins/genes [LOC109953912 and calcium Binding Protein 39 Like (cab39l)]. Notably, we detected significantly higher levels of Cu2+ during the sex change process, suggesting that Cu2+ is a male-related metal metabolite that may have an important function in male reproductive development.ConclusionsIn summary, we analyzed the protein profiles of ricefield eel gonadal tissues in five sexual stages (OV, IE, IM, IL, and TE) and verified the plausibility of the data. After preforming the functional enrichment of metal metabolite-related DEPs, we detected the contents of the metal metabolites Zn2+, Cu2+, Ca2+, and Fe2+/Fe3+ at these five stages and screened for (Cu-Zn) Sod1 and Mmp-9 as possible key proteins in the sex reversal process.

Effect of the combination of fluoxymesterone and a commercial blend of additives on the sex proportion, growth, and survival of Nile tilapia (Oreochromis niloticus) fry

In the commercial culture of Nile tilapia (Oreochromis niloticus), monosex, all-male populations obtained through steroid sex reversal is the most commercially used method to eliminate unwanted reproduction and increase final productive and economic yield. However, the rapid growth of Nile tilapia culture has led, in some regions, to an increase in infectious diseases and mortality in all stages of commercial culture. To reduce this, without the abuse of antibiotics or similar substances, feed additives are currently used in all stages, including the fry period, during the sex reversal process. The objective of this work was to evaluate the effect of synthetic steroid Fluoxymesterone (FM) at various concentrations in combination with a commercial Blend of additives (BA) on the percentage of males, growth, and survival of Nile tilapia fry. Five treatments (FM20, FM5+BA, FM10+BA, FM20+BA, and BA+FM20) and a control group were evaluated. Treatments differ in the order in which the commercial blend (2 g) was added and the FM concentration (5, 10, and 20 mg). The feeding trial lasted for 30 days using a sexually undifferentiated population of Nile tilapia. The results showed a decrease in the percentage of males in all treatments supplemented with the BA, compared to the FM20 treatment (containing 20 mg without BA). Growth and survival were positively affected by the inclusion of the BA in all treatments, showing higher values than those observed in the control group. Further studies are necessary to determine the FM-BA interaction.

Effect ofhigh temperatures onsex ratio anddifferential expression analysis (RNA-seq) ofsex-determining genes inOreochromis niloticus from different river basins inBenin.

The high temperature sex reversal process leading to functional phenotypic masculinization during development has been widely described in Nile tilapia (Oreochromis n iloticus) under laboratory or aquaculture conditions and in the wild. In this study, we selected five wild populations of O. niloticus from different river basins in Benin and produced twenty full-sib families of mixed-sex (XY and XX) by natural reproduction. Progenies were exposed to room temperature or high (36.5°C) temperatures between 10 and 30 days post-fertilization (dpf). In control groups, we observed sex ratios from 40% to 60% males as expected, except for 3 families from the Gobé region which showed a bias towards males. High temperature treatment significantly increased male rates in each family up to 88%. Transcriptome analysis was performed by RNA-sequencing (RNA-seq) on brains and gonads from control and treated batches of six families at 15 dpf and 40 dpf. Analysis of differentially expressed genes, differentially spliced genes, and correlations with sex reversal was performed. In 40 dpf gonads, genes involved in sex determination such as dmrt1, cyp11c1, amh, cyp19a1b, ara, and dax1 were upregulated. In 15 dpf brains, a negative correlation was found between the expression of cyp19a1b and the reversal rate, while at 40 dpf a negative correlation was found between the expression of foxl2, cyp11c1, and sf1 and positive correlation was found between dmrt1 expression and reversal rate. Ontology analysis of the genes affected by high temperatures revealed that male sex differentiation processes, primary male sexual characteristics, autophagy, and cilium organization were affected. Based on these results, we conclude that sex reversal by high temperature treatment leads to similar modifications of the transcriptomes in the gonads and brains in offspring of different natural populations of Nile tilapia, which thus may activate a common cascade of reactions inducing sex reversal in progenies.

Transcriptomic profiling and novel insights into the effect of AG ablation on gonad development in Macrobrachium rosenbergii

Androgenic glands (AGs) regulate male sexual differentiation in M. rosenbergii, and AG ablation may induce sex-reversal and male-to-neofemale transformation. However, information on gonad development following AG ablation is scarce. To investigate the effects of gonad development and sex reversal after AG ablation, we utilized transcriptomic profiling, RT–qPCR, and histological and morphological observation. Transcriptomic profiling generated 42.32–42.92 million clean reads. A total of 94,747 unigenes were assembled with a total length, average length, N50, and GC content of 160,460,655 bp, 1693 bp, 3438 bp, and 39.86%, respectively. Hsp70, IGFBP7, Mar-Mrr, Serpin, TUBA3, CREB, EF1a, and BMP7 were identified as differentially expressed by gonad transcriptome profiling after AG ablation. Interestingly, the Rap1, Hippo, PI3K, oxytocin, thyroid hormone, and apoptosis signalling pathways were all found to be involved in the sex-reversal process by regulating cell proliferation and gonad development and maintaining homeostasis. Sexual morphology features and histological changes following AG ablation validated the sex reversal from male to neofemale. Sexual manipulation permits monosex culture while considerably improving output yield. Taken together, these findings provide insight into the sexual reversal mechanism of M. rosenbergii after AG ablation. The findings in this study may aid in the future investigation of the sex-reversal mechanisms in other decapod species and offer a new breeding and culture strategy for M. rosenbergii aquaculture industry.

Molecular and morphological changes in Nile tilapia ( Oreochromis niloticus ) gonads during high‐temperature‐induced masculinization

Genetic sex of Nile tilapia (Oreochromis niloticus) is determined by genotype (genetic sex determination). However, these fish are temperature-sensitive, and high-temperature-induced sex reversal is a common practice in research and tilapia farming. In this study, we characterized the molecular and morphological sex differentiation of Nile tilapia during the process of sex reversal induced by high temperatures. The offspring of parental pairs of XX and XY Nile tilapia underwent high-temperature treatment for 20 d beginning at 5 dpf (days post fertilization). Gonads were isolated from Nile tilapia at seven time points between 10–180 dpf. The histology results showed that gonads of high-temperature-induced XX pseudomales exhibited morphological differences from XX females during 25–35 dpf. qRT-PCR analysis revealed that the expression levels of cyp19a1a and foxl2 were temperature-sensitive, and were lowly expressed in the gonads of pseudomales from 35 dpf onward. Such findings were also consistent with the histological observations of gonadal morphological differentiation. The expression of the TGFβ superfamily member, gsdf, was increased in the gonads of pseudomales from 12 dpf onward. The transcription of kdm6bb was elevated by high-temperature treatment at 10–25 dpf and displayed sexual dimorphism from 35 dpf onward. Dmrt1 was specifically expressed in phenotypic males from 12 dpf onward. Collectively, this study identified the roles of specific genes involved in the process of high-temperature-induced masculinization in Nile tilapia and established robust molecular markers of phenotypic sex in Nile tilapia that will be useful for ongoing research related to sex control.

The Effect of Immersion Doses of Hybrid Coconut Water on Guppy (Poecillia reticulata) to Increase the Male Larvae Percentage

Abstract Guppy fish is one type of ornamental fish that has its charm where it has beautiful colours and a slender body shape. Male guppy fish are more attractive, have more beautiful colour patterns, longer tail fins, and a slimmer body than female guppies. Hence, their selling price is much higher and is in demand in the market in the ornamental fish industry. One way that can be done to produce male guppy fish is by using the sex reversal or masculinization method. The sex reversal method that can be used is to use a solution of young hybrid coconut water (Cocos nucifera) with different doses. The immersion process will affect the percentage of male sex in guppy fish because in the young coconut water solution there is a content of potassium which plays a role in the sex reversal process which functions to regulate testosterone regulation in the body and direct the work of androgens. The purpose of this study was to determine the effect of young coconut water on the sex ratio of male guppies resulting from the immersion of female broodfish and to determine the best dose for the success rate of masculinization of guppy fish. The research was conducted from April to June 2020 in guppy fish farmers in Sidomulyo Village, East Ungaran District, Semarang Regency. The test animal used was the Black Moscow fish. This study used an experimental method with a completely randomized design of 4 treatments and 3 replications. Treatment of different immersion doses of young coconut water solution used were A (without immersion), B (40%), C (45%), D (50%). The data observed included the percentage of the male sex, survival rate and water quality. The results obtained from the research showed that the percentage value of male guppy fish was treated A (50.57 ± 3.48%), B (84.90 ± 2.55%), C (90.42 ± 1.67%), D (96.90 ± 1.34%). The results of the survival rate are treatment A (98.91 ± 0.95%), B (95.58 ± 2.48%), C (91.25 ± 3.46%), D 84.36 ± 3.62 %. The results of water quality observations were in the form of temperatures ranging from 26.9-28.9 ℃, pH (7.2-7.9), and DO ranging from 3.5-5.5 mg/ L. The conclusion was that immersion of young coconut water solution in pregnant guppy broods with different doses had a significant effect (P <0.05) on the percentage of the male sex. The best treatment was treatment D (50%) which resulted in the percentage value of male sex 96.90 ± 1.34%. Keywords: Black Moscow, potassium, masculinization, androgen hormone

Identification of Histone Modifications Reveals a Role of H2b Monoubiquitination in Transcriptional Regulation of dmrt1 in Monopterus albus

Gonadal trans-differentiation from ovary to testis occurs in a same individual, suggesting a role of epigenetic regulation. However, histone modifications concerning the sex reversal process remain elusive. We analyzed histone modifications using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Chromatin immunoprecipitation followed by sequencing (ChIP-seq) technology was used to test chromatin immunoprecipitation of gonads. Western blot analysis was performed to analyze protein expression. Immunofluorescence analysis was conducted to localize proteins in gonadal tissues. Here, we report a developmental atlas of histone modifications in the gonadal differentiation, including acetylation, methylation, and ubiquitination. We provided a detail distribution map of these modification sites including novel histone modifications along histones H2a, H2b, H3, and H4, and revealed their relationship with types of gonadal differentiation. We then determined a testis-enriched histone modification site, H2b monoubiquitination at K120, and its association with spermatogenesis. ChIP-seq demonstrated that the modification was highly enriched in the male sex-determining gene dmrt1 (doublesex and mab-3 related transcription factor 1), in particular, in its exon regions, suggesting its role in transcriptional regulation of dmrt1 in testis. Together, these data not only provide a new resource for epigenetic study in gonadal development, but also define an association of histone modifications with gonadal differentiation from ovary to testis.

Characterization of Methyltestosterone Degrading Bacteria Isolated from Tilapia Masculinizing Ponds: Metabolic Intermediate, Glucose Amendments Effects, and Other Hormones Transformation

17α-Methyltestosterone (MT) is widely used synthetic androgenic steroid in the tilapia aquaculture industry for masculinization: a sex reversal process in which hormones are utilized to induce production of male fish. Although MT is beneficial for aquaculture, release of residual MT can cause adverse effects on wild organisms. The aims of this study were to identify MT-degrading bacteria and to characterize their degradation abilities under the conditions experienced in the environment. Nocardioides nitrophenolicus S303, Acinetobacter radioresistens B051, and Ochrobactrum haematophilum B052 were the most efficient MT-degrading bacterial strains, with the shortest degradation half-life of 10–70 h. The MT degradation by Acinetobacter and Ochrobactrum has not been reported before. After comparing their degradation rates and for reason of biosafety, N. nitrophenolicus S303 was selected for further study. Although this strain degraded MT and testosterones, it could not degrade estrogens (estrone, 17β-estradiol, nor 17α-ethinylestradiol). Glucose amendment did not affect the MT degradation rate. No metabolites with androgenic activity were observed after 264-h treatment with this strain under aerobic conditions. Methandrostenolone was found as the major intermediate during 39–90 h. This is the first report indicating the 1,2-dehydrogenase activity in steroid clevage in N. nitrophenolicus. Our study provides important information concerning the application of N. nitrophenolicus S303 to enhance MT degradation in the environment.

Ocean acidification influences plant-animal interactions: The effect of Cocconeis scutellum parva on the sex reversal of Hippolyte inermis.

Ocean acidification (O.A.) influences the ecology of oceans and it may impact plant-animal interactions at various levels. Seagrass meadows located at acidified vents in the Bay of Naples (Italy) are considered an open window to forecast the effects of global-changes on aquatic communities. Epiphytic diatoms of the genus Cocconeis are abundant in seagrass meadows, including acidified environments, where they play key ecological roles. A still-unknown apoptogenic compound produced by Cocconeis triggers the suicide of the androgenic gland of Hippolyte inermis Leach 1816, a protandric hermaphroditic shrimp distributed in P. oceanica meadows located both at normal pH and in acidified vents. Feeding on Cocconeis sp. was proven important for the stability of the shrimp’s natural populations. Since O.A. affects the physiology of diatoms, we investigated if, in future scenarios of O.A., Cocconeis scutellum parva will still produce an effect on shrimp’s physiology. Cell densities of Cocconeis scutellum parva cultivated in custom-designed photobioreactors at two pH conditions (pH 7.7 and 8.2) were compared. In addition, we determined the effects of the ingestion of diatoms on the process of sex reversal of H. inermis and we calculated the % female on the total of mature individuals-1 (F/mat). We observed significant differences in cell densities of C. scutellum parva at the two pH conditions. In fact, the highest cell densities (148,808 ±13,935 cells. mm-2) was obtained at day 13 (pH 7.7) and it is higher than the highest cell densities (38,066 (±4,166) cells. mm-2, day 13) produced at pH 8.2. Diatoms cultured at acidified conditions changed their metabolism. In fact, diatoms grown in acidified conditions produced in H. inermis a proportion of females (F/mat 36.3 ±5.9%) significantly lower than diatoms produced at normal pH (68.5 ±2.8), and it was not significantly different from that elicited by negative controls (31.7 ±5.6%).

Socially controlled male-to-female sex reversal in the protogynous orange-spotted grouper, Epinephelus coioides.

Socially controlled sex change in teleosts is a dramatic example of adaptive reproductive plasticity. In many cases, the occurrence of sex change is triggered by a change in the social context, such as the disappearance of the dominant individual. The orange-spotted grouper Epinephelus coioides is a typical protogynous hermaphrodite fish that changes sex from female to male and remains male throughout its life span. In this study, male-to-female sex reversal in male Epinephelus coioides was successfully induced by social isolation. The body length and mass, gonadal change, serum sex steroid hormone levels and sex-related gene expression patterns during the process of socially controlled male-to-female sex reversal in E. coioides were systematically examined. This report investigates the physiological mechanisms of the socially controlled male-to-female sex reversal process in a protogynous hermaphrodite grouper species. The results enable us to study the physiological control of sex change, not only from female to male, but also from male to female.

MT-Feeding-Induced Impermanent Sex Reversal in the Orange-Spotted Grouper during Sex Differentiation.

In this study, we systematically investigated the process of sex reversal induced by 17-methyltestosterone (MT) feeding and MT-feeding withdrawal at the ovary differentiation stage in orange-spotted groupers, Epinephelus coioides. Gonadal histology showed that MT feeding induced a precocious sex reversal from immature ovaries to testes, bypassing the formation of an ovarian cavity, and MT-feeding withdrawal led to an ovarian fate. In both the MT feeding and MT-feeding withdrawal phases, cytochrome P450 family 11 subfamily B (cyp11b) gene expression and serum 11-KT levels were not significantly changed, suggesting that the MT-treated fish did not generate endogenous steroids, even though active spermatogenesis occurred. Finally, by tracing doublesex-expressing and Mab-3-related transcription factor 1 (dmrt1)-expressing cells and TUNEL (terminal deoxynucleotidyl transferase 2-deoxyuridine, 5-triphosphate nick end labeling) assays, we found that the efferent duct formed first, and then, the germ cells and somatic cells of the testicular tissue were generated around the efferent duct during MT-feeding-induced precocious sex reversal. Collectively, our findings provide insights into the molecular and cellular mechanisms underlying sex reversal induced by exogenous hormones during sex differentiation in the protogynous orange-spotted grouper.

The sex change of the caridean shrimp Hippolyte inermis Leach: temporal development of the gonopore morphology

Sex reversal is a process observed in several marine organisms, including some lineages of caridean shrimps. We investigated the gonopore shape and size, to study the sex reversal using Hippolyte inermis as a model. A method was developed which can be applied to identify the sex in juveniles of H. inermis, especially, useful when the standard method of sex assessment is not applicable. The position and the shape of gonopores was recorded under a light macroscope. The sex of mature individuals was then determined by observing the presence/absence of the appendix masculina. In addition, analysis of ontogenetic changes of gonopores were performed to compare their morphology with other species of shrimps whose gonopore morphology was previously known. Female gonopores are located at the far proximo-medial end of the third pair of pereiopod coxae and distally they bear cup-shaped structures, whilst male gonopores are located at the far proximo-medial end of the fifth pair of pereiopod coxae and they have a simpler structure. The shape and structure of gonopores in H. inermis resembled that of other caridean decapods. Intersex individuals were never observed, although this species was demonstrated to be protandric. This observation confirmed previous assumptions indicating that the process of sex reversal is very fast in H. inermis and that it takes place within a single moult. The identification of sex based on the position and shape of gonopores is feasible in this species, and it provides helpful insights for studying sex reversal in small decapods.

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Seasonal expression of arginine vasotocin mRNA and its correlations to gonadal steroidogenic enzymes and sexually dimorphic coloration during sex reversal in the gilthead seabream (Sparus aurata).

Arginine vasotocin is a hormone produced in the hypothalamus of teleost fish that has been shown to regulate gonad development and sexual behavior. To study the role of arginine vasotocin in the gonadal cycle of the hermaphrodite gilthead seabream, Sparus aurata, we cloned the seabream arginine vasotocin (avt) complementary DNA (cDNA). We investigated the expression of brain avt throughout the gonad cycle using real-time quantitative PCR and compared its expression levels to the expression levels of two key gonadal steroidogenic enzymes, cyp19a1a and cyp11b2. In July, when the process of sex reversal is thought to begin, avt expression was elevated over the previous 2months. Avt in the brain remained at or above the level of July until November then peaked again in December. There was no difference between males and females in the expression levels of brain avt throughout the year. However, only in ambisexual fish was the expression of the cyp19a1a gonadal aromatase correlated to the expression of avt in the brain. Cyp11b2 did not show any correlation to brain avt expression. We also found that females had more intense body coloration than males and that this intensity peaked prior to spawning. Avt expression and female coloration were positively correlated. The fact that brain avt expression was lowest during gonad quiescence, together with the observation of a correlation between brain avt with gonadal cyp19a1a and body coloration during that time suggests that avt may play a role during the process of sex reversal and spawning of the gilthead seabream.

Progress in studies of sex determination mechanisms and sex control techniques in Cynoglossus semilaevis (half-smooth tongue sole)

The Cynoglossus semilaevis (half-smooth tongue sole) is a marine flatfish of great commercial value for fisheries and aquaculture in China. It has a female heterogametic sex determination system (ZW/ZZ) and environmental factors can induce sex-reversal of females to phenotypic males, suggesting that it is a promising model for the study of sex determination mechanisms. Additionally, females grow much faster than males and it is feasible to improve the aquaculture production through sex control techniques. This paper reviews the progress in research on sex determination mechanisms research in our laboratory. We have completed whole-genome sequencing and revealed the genome organization and sex chromosome evolution of C. semilaevis. A putative male determining gene dmrt1 was identified and DNA methylation was verified as having a crucial role in the sex reversal process. Genetic maps and sex-specific biomarkers have been used in a marker-assisted selection breeding program and for differentiation of the fish sex. Development and improvement of sex control technologies, including artificial gynogenesis and production of breeding fry with high proportion of females, is also reviewed. These research advances have provided insight into the regulation of sex determination and enabled efficient sex management in artificial culturing of C. semilaevis.

High-throughput sequencing analyses of XX genital ridges lacking FOXL2 reveal DMRT1 up-regulation before SOX9 expression during the sex-reversal process in goats.

FOXL2 loss of function in goats leads to the early transdifferentiation of ovaries into testes, then to the full sex reversal of XX homozygous mutants. By contrast, Foxl2 loss of function in mice induces an arrest of follicle formation after birth, followed by complete female sterility. In order to understand the molecular role of FOXL2 during ovarian differentiation in the goat species, putative FOXL2 target genes were determined at the earliest stage of gonadal sex-specific differentiation by comparing the mRNA profiles of XX gonads expressing the FOXL2 protein or not. Of these 163 deregulated genes, around two-thirds corresponded to testicular genes that were up-regulated when FOXL2 was absent, and only 19 represented female-associated genes, down-regulated in the absence of FOXL2. FOXL2 should therefore be viewed as an antitestis gene rather than as a female-promoting gene. In particular, the key testis-determining gene DMRT1 was found to be up-regulated ahead of SOX9, thus suggesting in goats that SOX9 primary up-regulation may require DMRT1. Overall, our results equated to FOXL2 being an antitestis gene, allowing us to propose an alternative model for the sex-determination process in goats that differs slightly from that demonstrated in mice.

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  This review attempts to group the recent hypotheses involved in the complex system of determination and sex differentiation in fish. Based on recent literature, we relate the key genes involved in the genomic cascade as the Cyp19, Dmrt1, Sox9, Foxl2, Esr, Dax1, Sf1 and Amh1, and still little known action of temperature on them. As the sex reversal is a highly desired process in fish farming aiming at obtaining male mono-sexual populations (due to weight gain of males), several techniques based on direct and indirect manipulation of phenotypic sex are being tested. Recent surveys show the use of temperature as alternative to the process of sex reversal. However, high or low temperatures have limited effect, in addition to there being a window of sex reversal in which temperature acts, varying from species to species. Thus, we draw a parallel with the role of temperature in the process of sex reversal and its effect on genes of the genomic cascade, which has been the subject of several studies that attempt to explain how temperature would be acting in this process. Intracellular receptors, such as those used for steroid hormones, act as transcription factors to regulate target genes moving between the nucleus and cytoplasm and, in the hormone absence, are linked to the complex of heat shock protein 90 kDa (Hsp90). Through this mechanism, it is possible to predict that fluctuations in temperature can influence the action of hormones, the increased transcription of genes involved in steroidogenesis and hence in sexual differentiation, becoming an alternative to explain where the temperature is acting. However, this literature review discusses the correlations between the genomic cascade, the action of intracellular receptors and the influence of temperature within this large system of determination and sex differentiation in fish.   Key words: Fish, gene, sex differentiation, temperature.

Sexual patterns and protogynous sex reversal in the rusty parrotfish, Scarus ferrugineus (Scaridae): histological and physiological studies

Gonadal histology confirmed that Scarus ferrugineus is a diandric protogynous fish. The process of protogynous sex reversal was investigated through histological observations on the gonads of females changing sex to male. This process was divided into three stages on the basis of changes in the structure of the germinal and somatic elements. Ovaries of functional females (stages IV-V) were filled with vitellogenic oocytes during the breeding season but contained no trace of spermatogenic tissue. During post-spawning period, the remaining vitellogenic oocytes began to degenerate and accompanied by a drop in plasma levels of estradiol-17β. At the commencement of sex change, previtellogenic oocytes began to degenerate and stromal cell aggregation was observed in the central region of the lamellae. At mid-reversal stage, steroid-producing cells (Leydig cells) developed at the border of the stromal aggregate and spermatogonial cysts appear at the periphery of lamellae. Finally, sex change to secondary males was considered complete, with the beginning of active spermatogenesis and spermiation. Plasma levels of testosterone remained low throughout the sex change, but II-KT increased rapidly parallel to the increased number of Leydig cells while the level of estradiol-17β decreased. The results indicate also that the sex-changed males had higher level of II-KT than primary males, while primary males had higher level of testosterone. Histological examination revealed that testes of primary and secondary males are almost identical in organization of the spermatogenic cysts, association of sertoli cells, and developing germ cells but differ in clustering and development of Leydig cells.

Effects of Estradiol Valerate on Steroid Hormones and Sex Reversal of Female Rice Field Eel, Monopterus albus (Zuiew)

The rice field eel, Monopterus albus (Zuiew), is an important aquaculture fish in many Asian countries. Productivity, however, is limited by low fecundity as a result of sex reversal of female juveniles of this fish. This study was conducted to investigate the effects of estradiol valerate on the sex reversal process of this fish by determining sex changes of initial 2-yr-old female eels (26.5 ± 2.6 cm, 21.9 ± 2.1 g) treated with three concentrations of estradiol valerate: 0 mg/kg (Control, Group A), 10 mg/kg (Group B), and 50 mg/kg (Group C). In Group A, sex reversal took place with significantly higher percentage of sex change: about 45% of female eels to intersexes and males. Only about 22% of female eels changed to intersexes and males in Group B, but no sex reversal occurred in Group C during the experimental period. The result also indicated that the gonadal development and the concentrations of serum estradiol (E2) and testosterone (T) varied significantly with different concentrations of dietary estradiol valerate. Serum T significantly reduced but serum E2 significantly increased in Group C compared to the control Group A. The results demonstrated that estradiol valerate affected the secretion of sex steroid hormones in rice field eels and thus inhibited the sex reversal process at a low concentration (10 mg/kg) or stopped the sex reversal process at a high concentration (50 mg/kg) of estradiol valerate. These results suggest that a higher yield of mature female eels could be achieved by treatments with estradiol valerate, thus enhancing the fecundity of this rice field eel.

Decontamination trends in the aquacultured fish gilthead seabream ( Sparusaurata) after feeding long-term a PCDD/F spiked feed

Aquacultured fish gilthead seabream ( Sparus aurata), previously exposed to low levels of polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/Fs) trough the diet for a 13 months period, were fed on a clean feed for another 22 months. Gilthead seabream is a protandrous hermaphrodite species and this “decontamination” period coincided with the stages of sex differentiation, maturity and reproduction of the fish. PCDD/F levels in the fish tissues (i.e. muscle, liver, perivisceral fat and gonads), expressed in pg WHO-TEQ g −1 fresh weight, showed a general decreasing trend during the “decontamination” period. However, this general trend varied among tissues and was also dependent on sex and lipid contents. Toxicological effects affecting fish behaviour and hepatic marker responses were also evaluated. The results pointed out that exposure to PCDD/Fs did not have an impact on fish development and reproduction, since the proportion of sexes found after the sex reversal process was within the normal range described for this species. In addition, long-term exposure to low PCDD/F levels did not significantly affect the response of most of the biochemical markers considered. On the contrary, some of them (e.g. EROD activity) showed variations in their responses during the sex differentiation process and onwards. Finally, the hepatic AhR mRNA levels increased during dioxin exposure but they returned to values typical for non-exposed fish after the “decontamination” period.