Gonadal restructuring and correlative steroid hormone profiles during natural sex change in protogynous honeycomb grouper (Epinephelus merra)

The honeycomb grouper shows protogynous hermaphroditism. The endocrine mechanisms involved in gonadal restructuring throughout protogynous sex change are largely unknown. In the present study, we investigated changes in the gonadal structures and levels of serum sex steroid hormones during female to male sex change in the honeycomb grouper. On the basis of histological changes, entire process of sex change was assigned into four developmental phases: female, early transition (ET), late transition (LT), and male phase. At the female phase, the oocytes of several developmental stages were observed including gonial germ cells in the periphery of ovigerous lamellae. At the beginning of ET phase, perinucleolar and previtellogenic oocytes began degenerating, followed by proliferation of spermatogonia toward the center of lamella. The LT phase was characterized by further degeneration of oocytes and rapid proliferation of spermatogenic germ cells throughout the gonad. At the male phase, no ovarian cells were observed and testis had germ cells undergoing active spermatogenesis. Serum levels of estradiol-17beta (E2) were high in females in the breeding season, but low in the non-breeding female, transitional and male phase, and those of 11-ketotestosterone (11-KT) and testosterone (T) were low in females and gradually increased in the transitional and male phase. The present results suggest that low serum E2 levels and degeneration of oocytes accompanied by concomitant increase in the 11-KT levels and proliferation of spermatogenic germ cells are probably the events mediating protogynous sex change in the honeycomb grouper.     

Protogynous hermaphroditism in the half-banded sea perch, Hypoplectrodes maccullochi (Serranidae)

The half-banded sea perch, Hypoplectrodes maccullochi (Serranidae) exhibits protogynous hermaphroditism, based on the presence of transitional individuals collected from rocky reefs along the coast of New South Wales, Australia. Males dominated the larger size and age classes, whereas females predominated the smaller size and age groups. Histological preparations suggested that sex change occurs between 55–85 mm s.l. and 1 + to 4+ years of age. Three transitional phases were identified which varied according to the proportion of gametogenic tissue and remnant oocytes present within their gonads. All individuals greater than 85 mm s.l. were functional males containing well defined crypts of spermatids and spermatozoa. Remnant oocytes within these testes indicated prior female function. H. maccullochi were sexually monochromatic. The occurrence of some small males derived from pre-spawning females suggested pre-maturational sex change occurred in some individuals. Sex changing individuals were found in all the months of the year that gonads were examined in detail (January, February, April and June 1989). Fish were ripe in August and November. H. maccullochi conforms with the pattern of reproduction evident within the genus Hypopleclrodes .     

Seasonal cycles of gonadal development and plasma sex steroid levels in Epinephelus morio, a protogynous grouper in the eastern Gulf of Mexico

In a field population of the protogynous red grouper Epinephelus morio in the eastern Gulf of Mexico, females with oocytes at all stages of development were collected during the spawning season suggesting that several batches of oocytes may be released over the spawning period. Plasma oestradiol (E₂) levels were highest in ripe females whose gonads contained both cortical alveoli and vitellogenic oocytes during the breeding season. Males were still spermiating as late as August, although levels of androgens 11-ketotestosterone (11-KT) and testosterone (T) had declined from their peaks in March. A few red grouper with either perinucleolar or cortical alveoli stage oocytes were undergoing sex change both during and after the spawning period. Low levels of E₂, T and 11-KT were detected in transitionals. Proliferation of male tissue was not restricted to any specific area of the gonad but occurred in pockets within the ovarian lumen. The sequence of an increase in gonial cells along the periphery of the lamellae, increase in interstitial tissue, degradation of female elements, and formation of a sperm duct seemed to be concurrent with spermatocyte proliferation and the process of preparing the gonad to function as a testis.     

Anatomical changes to the gonad during protogynous sex change in the half-moon grouper Epinephelus rivulatus (Valenciennes)

Anatomical changes to the gonad during sex change in the protogynous grouper Epinephelus rivulatus are described from histological observations. A decrease in ovarian mass occurred soon after the onset of sex change as oocytes atrophied and were removed from the gonad. Blood supply and abundance of unidentified somatic cells increased at this time as proliferation of sperm tissue commenced. As the gonad was cleared of ovarian tissue, the rate of spermatogenesis increased and the lamellae soon became dominated by sperm and connective tissue. Putative Leydig cells, the probable sites of male steroid production, appeared in transitional gonads and were most abundant in the testes of immature males. Peripheral sperm sinuses subsequently formed within basal tissue layers of the tunica and expanded as they filled with spermatids. The process of sex change, occurring as a result of experimental manipulation of wild populations at the start of the spawning season, took c. 3 weeks. This appears rapid compared to other hermaphroditic species and may reduce the impacts of fishing on reproductive output by E. rivulatus populations.     

Sexual differentiation,gonad development,and spawning seasonality of the Hawaiian butterflyfish,Chaetodon multicinctus

Synopsis The reproductive biology of the coral reef butterflyfish,Chaetodon multicinctus, was investigated by histological examination of gonads sampled over an 18 month period from a shallow inshore population on Oahu, Hawaii. Most gonads developed directly from previously undifferentiated tissue. Ovarian development (the structural formation of lamellae and primary oocytes) was observed in fish ≥44 mm and testicular development (the formation of spermatogenic crypts) in fish ≥62 mm standard length (SL). In addition, testis formation was identified within the ovarian lamellae of several differentiated but immature fish. It is hypothesized that prematurational sex change may facilitate monogamy within the highly competitive social structure of this site attached species. Oocyte development in mature females was marked by distinct phases of primary growth, the formation of yolk vesicles, and vitellogenesis. Spawning activity was histologically identified by the maturation and hydration of fully yolked oocytes, and presence of postovulatory follicles. Recently spawned females from field collections and experimental gonadotropin-treatments exhibited postovulatory follicles that were estimated to persist at least 24 h after ovulation. Atresia of yolked oocytes was classified into four stages of cell degeneration and resorption. Monthly analyses of oocyte development and atresia within the sample population show thatC. multicinctus has a protracted annual spawning season with a major peak during the early spring and evidence of spawning activity among some individuals in the fall. Histological analyses of spawning activity provide more accurate and unambiguous information than do traditional gonadosomatic assays in this and probably other coral reef fishes.     

Histological and ultrastructural evidence for the role of gonadal steroid hormones in sex change in the protogynous wrasse Thalassoma duperrey

Synopsis The process of sex change in the protogynous wrasse, Thalassoma duperrey, was investigated through histological and ultrastructural observations on the gonads of females changing sex to male. Changes in plasma steroid levels concomitant with structural changes were measured by radioimmunoassay. The process of sex change from ovary to testis was divided into six stages on the basis of changes in the structure of the germinal and somatic elements. Ovaries of females were filled with vitellogenic oocytes during the breeding season, but contained no spermatogenic tissue (Stage 1). At the commencement of sex change (Stage 2), vitellogenic oocytes began to degenerate, and were ingested by macrophagous cells. This stage was accompanied by a rapid drop in plasma levels of estradiol-17. Thereafter, previtellogenic oocytes (Stage 3) also began to degenerate, and aggregations of stromal tissue, and loose connective tissue were observed in the central region of the lamellae. Steroid producing cells (Leydig cells), developed at the border of this loose connective tissue. Presumed spermatogonia proliferated on the periphery of the lamellae, and Leydig cells increased in size and number (Stage 4). Spermatogonia formed cysts, and underwent spermatogenesis (Stage 5). Finally, sex change to male was considered complete, with the beginning of active spermatogenesis and spermiation (Stage 6). Plasma levels of testosterone remained low throughout the sex change, but a second androgen, 11-ketotestosterone increased gradually in parallel to the increased numbers of Leydig cells and spermatogonia. Preliminary in vitro incubation of gonads with salmon gonadotropin, revealed that sex-changed males had higher levels of 11-ketotestosterone production than did females, while females had higher levels of estradiol-17 production than did males. Production of both these steroids increased in a dose-related fashion with increasing doses of gonadotropin.     

Gonadal development and non-functional protogyny in a coral-reef damselfish, Dascyllus albisella Gill

All gonads of the Hawaiian dascyllus Dascyllus albisella , irrespective of the final sex of individuals, developed an ovarian lumen and primary-growth-stage oocytes after an initially undifferentiated state. From this ovarian state or from more differentiated ovaries, some gonads redifferentiated into testes. None of 117 individuals examined had a gonad containing degenerating vitellogenic oocytes and proliferating spermatogenic tissue. Eleven individuals had gonads containing degenerating cortical-alveolus-stage oocytes and developing spermatogenic tissue. The size of these individuals overlapped with the female size range in which the majority of the females were still in the middle of the maturation process. They were absent from the larger size range where the majority of females had vitellogenic oocytes. This indicated that the transition toward maleness is likely to have occurred after the onset of cortical-alveolus stage, but before final oocyte maturation and spawning as females. Therefore the protogynous pattern of gonadal development was non-functional. There was no dimorphism in the sperm duct configuration, and all the testes were secondary testes reported for diandric, protogynous species with undelimited gonads. Very early development of an ovarian lumen appeared to have resulted in a secondary-male configuration in all testes, although redifferentiation into males appeared to have occurred before sexual maturity and spawning as females.     

Gonadal development and diandric protogyny in two populations of Dascyllus reticulatus from Madang, Papua New Guinea

Two Dascyllus reticulatus populations from Madang, Papua New Guinea exhibited diandric protogyny. In both populations, gonads began as undifferentiated, and then developed oocytes in the primary growth stage and an ovarian lumen. From this ovarian state or from more developed ovaries containing oocytes beyond the primary‐growth stage, some gonads developed into testes. The first sign of testicular development was degeneration of oocytes, degeneration of oocytes in the primary growth stage in ovarian gonads and degeneration of oocytes of all growth stages present including the primary growth stage in ovaries, which was then followed by development of spermatogenic tissue. In both populations, most of the fish that had gonads with degenerating oocytes were smaller than the smallest mature females, indicating that development towards testes was mostly initiated in immature gonads containing only pre‐vitellogenic oocytes. On some occasions, however, females as large as other mature females also had gonads with degenerating oocytes, suggesting that development towards testes may have occurred in mature ovaries as well. This latter notion is further strengthened by the discovery of a fish having a gonad that contained both degenerating vitellogenic oocytes and developing spermatogenic tissue. Taken together, these results suggest that D. reticulatus can exhibit diandric protogyny, because testes in D. reticulatus developed from juvenile gonads as well as from mature ovaries.     

Gonadal development and an indication of functional protogyny in the Indian damselfish (Dascyllus carneus)

The objective of this study was to determine the sexual pattern of the Indian dascyllus Dascyllus carneus . After an initially undifferentiated state, gonads of D. carneus developed an ovarian lumen and primary growth stage oocytes, and subsequently cortical-alveolus stage oocytes. From ovaries with cortical-alveolus stage oocytes and from more developed ovaries, some gonads redifferentiated into testes. From a sample of 163 individuals, two had a gonad containing degenerating vitellogenic oocytes and proliferating spermatogenic tissue, nine had a gonad containing degenerating cortical-alveolus stage oocytes and spermatogenic tissue, and five had a gonad with degenerating primary growth stage oocytes and spermatogenic tissue. The size of these individuals overlapped greatly with the size range of mature females, suggesting that at least in some individuals, redifferentiation toward a testis occurred after spawning as females. This indicates that D. carneus is a functional, diandric protogynous hermaphrodite. Removal of a dominant male(s) did not induce a sex change in any of the ranking females in the laboratory and field groups. There was no difference in the number of chases and signal jumps performed by the ranking female between control and experimental field groups, or before and after removal of the male. However, the sizes of the ranking females were at or beyond the size range of individuals with a mixed-stage gonad, suggesting that the developmental window for female-to-male sex change may not be open ended. In 41 of 43 field groups, in which sex of fish was determined histologically or by the shape of the urogenital papilla, one to several highest size ranks were occupied by males, followed by one to several females. Mature males, however, were not limited to the highest ranks and occurred at various lower size ranks within groups. Individuals with a mixed-stage gonad also occupied various size ranks within groups.     

Early gonadal development and primary males in the protogynous epinepheline, Cephalopholis boenak

Early gonadal development of the protogynous epinepheline, Cephalopholis boenak, was examined histologically in 289 specimens with standard length (L_S) of 42–130 mm, collected from May 2000 to April 2002 in Hong Kong waters, to determine male developmental pathways and establish its sexual pattern. All juvenile gonads developed an ovarian lumen with primary‐growth stage oocytes and scattered spermatogenic tissue prior to sexual differentiation and first sexual maturation. From this bisexual phase containing both female and male tissues, some gonads differentiated as ovaries with further oocyte growth to cortical‐alveolus and vitellogenic stages, the rest differentiated as testes with the proliferation of spermatogenic tissue and the formation of a sperm sinus. All testes retained the lumen and primary‐growth stage oocytes, and sperm sinuses ran within the gonad wall. Unlike most protogynous species, among functional males it was impossible to distinguish those resulting from juveniles through sexual differentiation (i.e. primary male) from those resulting from functional females through sex change (i.e. secondary male) based solely on testicular morphology. A proportion‐spermatogenic‐tissue index (I_ST) was, therefore, developed and determined to be a reliable quantitative indicator for distinguishing differentiating, primary males before a sperm sinus was evident, from differentiating females during sexual differentiation. Since sexually transitional specimens with the concominant appearance of degenerating vitellogenic, or later, stage oocytes and spermatogenic tissue in the gonads were previously noted from Hong Kong, diandric, protogynous hermaphroditism is confirmed in C. boenak. For species, such as this and other epinephelines, in which all males have the same testicular morphology, a complete analysis of a wide range of body sizes from juveniles to adults is necessary for understanding male developmental pathways, and determining sexual pattern.     

Historical aspects of protandrous sex change in the anemonefish Amphiprion melanopus (Pomacentridae, Teleostei)

Gonadal structure and cellular composition were examined in juveniles, males and females of the protandric hermaphrodite, Amphiprion melanopus. Functional sex change was experimentally induced in the field and gonad structure was histologically examined both qualitatively and quantitatively at 10, 20, 30 and 45 days after its initiation. Juvenile gonads consist primarily of immature ovarian tissue. Functional male gonads are ovotestes with co-existing mature spermatogenic tissue and immature ovarian tissue, while females possess only ovarian tissue. The initiation of sex change is marked by a rapid maturation of spermatogenic tissue and proliferation of putative oogonia. Gonads were essentially female by 20 days into sex change, but evidence of mature female function (marked by the initiation of vitellogenesis) was not observed until 45 days. Considerable variation between individuals was seen in quantitative measures of gonadal change in the early stages of sex change, but not in later stages. Progress in sex change as indicated by histological indicators was, however, consistent within stages. Duet systems for gamete transport changed from the male to the female form after all male tissue had been replaced.     

Conserved form and function of the germinal epithelium through 500 million years of vertebrate evolution

The germinal epithelium, i.e., the site of germ cell production in males and females, has maintained a constant form and function throughout 500 million years of vertebrate evolution. The distinguishing characteristic of germinal epithelia among all vertebrates, males, and females, is the presence of germ cells among somatic epithelial cells. The somatic epithelial cells, Sertoli cells in males or follicle (granulosa) cells in females, encompass and isolate germ cells. Morphology of all vertebrate germinal epithelia conforms to the standard definition of an epithelium: epithelial cells are interconnected, border a body surface or lumen, are avascular and are supported by a basement membrane. Variation in morphology of gonads, which develop from the germinal epithelium, is correlated with the evolution of reproductive modes. In hagfishes, lampreys, and elasmobranchs, the germinal epithelia of males produce spermatocysts. A major rearrangement of testis morphology diagnoses osteichthyans: the spermatocysts are arranged in tubules or lobules. In protogynous (female to male) sex reversal in teleost fishes, female germinal epithelial cells (prefollicle cells) and oogonia transform into the first male somatic cells (Sertoli cells) and spermatogonia in the developing testis lobules. This common origin of cell types from the germinal epithelium in fishes with protogynous sex reversal supports the homology of Sertoli cells and follicle cells. Spermatogenesis in amphibians develops within spermatocysts in testis lobules. In amniotes vertebrates, the testis is composed of seminiferous tubules wherein spermatogenesis occurs radially. Emerging research indicates that some mammals do not have lifetime determinate fecundity. The fact emerged that germinal epithelia occur in the gonads of all vertebrates examined herein of both sexes and has the same form and function across all vertebrate taxa. Continued study of the form and function of the germinal epithelium in vertebrates will increasingly clarify our understanding of vertebrate reproduction. J. Morphol. 277:1014–1044, 2016. © 2016 Wiley Periodicals, Inc.     

Natural sex change in the temperate protogynous Ballan wrasse Labrus bergylta

Wild Ballan wrasse Labrus bergylta were sampled monthly over 2 years in western Norway to identify the natural process of sex change in this species. Light microscopy of standard histological‐stained and immunohistochemistry‐treated gonad tissue showed that spermatogonial germ cells tended to proliferate around the periphery of the lamellae before filling into the slowly receding, apoptotic central areas of the lamellae. Sex change occurred following the breeding season. From July to September, fish were most often in an early state of gonadal transition (ET), characterized by degenerating previtellogenic oocytes and pockets of proliferating spermatogonia in the germinative epithelia. The majority of fish with late transitional gonads, that were typically dominated by spermatogenic cells, developing efferent ducts and the beginning of lobule formation, were found between October and November. Sex steroid profiles of fish representing the different sexual phases showed that breeding females had the highest concentrations of 17β oestradiol (E₂) and the lowest concentration of 11 ketotestosterone (11KT). Concentrations of E₂ decreased greatly in ET fish at the beginning of sex change and remained low in all subsequent phases. The opposite trend was demonstrated in 11KT profiles. Initial‐phase female fish had minimal concentrations of 11KT, but these increased during subsequent transitions. Sex change occurred most often in fish 34–41 cm total length (L_T) and the median of fish in the size‐frequency overlap of female and male fish was 36 cm L_T.     

High‐density aggregations of rodlet cells in the gonads of Greenland halibut Reinhardtius hippoglossoides,a deep‐water marine flatfish

Large aggregations of rodlet cells in the gonads of male and female Greenland halibut Reinhardtius hippoglossoides are reported for the first time. These rodlet cells were not arranged epithelially but rather were found throughout the connective tissue between oocytes (females) or within lymphatic spaces between testicular lobules (males). The reason for large aggregations of rodlet cells in the gonads and not other tissues of this species is uncertain.     

Efferent duct differentiation during female-to-male sex change in honeycomb grouper Epinephelus merra

Efferent duct (ED) differentiation was examined histologically during female-to-male sex change in the honeycomb grouper Epinephelus merra . During natural sex change, ED differentiation began with the appearance of slit-like structures between the stromal tissue and the tunica ovary and small oval-shaped spaces within the wall of the ovarian cavity, accompanied by oocyte degeneration and the initiation of spermatogenesis, i.e. the early transitional phase. In the late transitional phase, ED structure formation expanded and further ED differentiation occurred, including the rapid multiplication of spermatogonial germ cells. In sex-changed males, the slit-like structures increased in size, fused with each other and finally formed a well-developed ED. The oval-shaped spaces also increased in size and fused to form an ED. In contrast, during artificial sex change, induced by aromatase inhibitor (AI, 1 mg kg⁻¹), ED differentiation in E. merra was first observed as the appearance of slit-like structures and small oval-shaped spaces in the restructuring gonads in the third week after AI treatment. These were accompanied by oocyte degeneration and the proliferation of gonial germ cells into spermatogonia. In the fifth week, the rapid multiplication of spermatogonial germ cells, increases in 11-ketotestosterone (11-KT), and further differentiation of EDs were observed. Sex-changed males had testes containing sperm in the completely differentiated EDs; the significantly highest levels of 11-KT were observed in the sixth week. Simultaneous increases in 11-KT and initiation of ED differentiation were observed, suggesting a role of 11-KT in ED differentiation during sex change. There were no basic differences in the mechanisms of natural and artificially induced ED differentiation. Two types of structure led to the formation of EDs in two different areas of the newly formed testis during sex change.     

PROCESS OF FUNCTIONAL SEX REVERSAL OF THE GONAD IN THE FEMALE GUPPY, POECILIA RETICULATA, TREATED WITH ANDROGEN BEFORE BIRTH

In the guppy, Poecilia reticulata , ovarian differentiation occurs during the embryonal life by 14 days after the preceding parturition. Testicular differentiation begins with the appearance of prominent aggregations of stroma cells in the gonadal hilus occurring by 18 days following the last parturition.
Oral administration of methyltestosterone (400 μ/g diet) to gravid guppies, begun 13–15 days after the preceding parturition and continued until the end of gestation, induced a male-type aggregation of somatic cells in the hilus of ovaries of female embryos. Gonads of newly born, androgenized females still had developing oocytes but were always provided with atypical clusters of stroma cells in their hilus. The gonads of affected female offspring developed successively into definite testes within 20 days after birth, displaying a precocious differentiation of the hilar stroma into sperm ducts and testicular interstitium, a concomitant initiation of spermatogenesis, and a conspicuous degeneration of oocytes. A successful masculinization of the somatic element, which may occur prior to that of the germ cells, in androgen-affected embryonic ovaries seems to be essential for the functional sex reversal of genetic females in the guppy.     

Histological and ultrastructural investigation of early gonad development and sex differentiation in Adriatic sturgeon (Acipenser naccarii,Acipenseriformes, Chondrostei)

Gonad development and sex differentiation from embryos to 594‐day‐old individuals were investigated in farmed Acipenser naccarii using light and transmission electron microscopy. The migrating primordial germ cells first appear along the dorsal wall of the body cavity in embryos 1.5 days before hatching. The gonadal ridge, containing a few primary primordial germ cells (PGC‐1) surrounded by enveloping cells, appears in 16‐day‐old larvae. At 60 days, the undifferentiated gonad is lamellar and PGC‐1 multiply, producing PGC‐2. In 105‐day‐old juveniles, a distinct germinal area with advanced PGC‐2 appears on the lateral side near the mesogonium and the first blood vessels are visible. At 180 days, putative ovaries with a notched gonadal epithelium and putative testes with a smooth one appear, together with adipose tissue on the distal side. In 210‐day‐old juveniles, active proliferation of germ cells begins in the putative ovaries, whereas putative testes still contain only a few germ cells. The onset of meiosis and reorganization of stromal tissue occurs in ovaries of 292‐day‐old individuals. Ovaries with developed lamellae enclosing early oocyte clusters and follicles with perinucleolar oocytes occur at 594 days. Meiotic stages are never found, even in anastomozing tubular testes of 594‐day‐old individuals. Steroid producing cells are detected in the undifferentiated gonad and in the differentiated ones of both sexes. Anatomical differentiation of the gonad precedes cytological differentiation and female differentiation largely precedes that of the male. Gonad development and differentiation are also associated with structural changes of connective tissue, viz. collagen‐rich areas are massive in developing testes and reduced in ovaries. J. Morphol., 2008. © 2008 Wiley‐Liss, Inc.     

Gonad morphology and sex succession in the protogynous hermaphrodite Hemanthias vivanus (Jordan and Swain)

Hemanthias vivanus , an anthiine serranid common on deep (70–80 m) reefs off eastcentral Florida, is shown to be a protogynous hermaphrodite with observed size ranges as follows: female, 49–77 mm S.L.; transitional 65–74; male, 65–96. Sexes are distinguishable externally as the dorsal spine filaments elongate and colour pattern changes with sex succession. In females, lamellae extend from the lateral and dorsal walls of the bilobed ovary into the lumen. Ova mature within the lamellae and at ovulation pass centrally into the lumen, posteriorly into a common post-ovarian sinus, and exit through the oviduct. Females possess a discrete tissue band (testicular islets) at the border of the alamellar region, from which testicular tissue proliferates during sex succession and replaces the regressing ovarian tissue. In males, spermatozoa are formed in crypts which drain peripherally into a sperm sinus which forms by rupture of muscular and connective tissue layers surrounding the gonad. The sperm sinus drains posteriorly into the vas deferens which is formed by rupture of collagenous tissue surrounding and posterior to the now vestigial post-ovarian sinus. The vas deferens and mesonephric duct join within the urogenital papilla. Two smooth muscle bands associated with the infracarinalis medius muscle bands, the collagenous tissue surrounding the gonoduct, and the intestinal lining are implicated in the extrusion of gametes, urine and faeces. At sex succession, ova are resorbed in one of at least three pathways depending upon their stage of development at the onset of sex transformation: unovulated mature ova become encysted with fibroblasts and invaded by cells of an unknown origin; non-mature yolky ova are invaded and phagocytized by granulosa cells; non-yolky ova are not invaded but fragment.     

Sex differentiation and aspects of gametogenesis in shortnose sturgeon Acipenser brevirostrum Lesueur

Shortnose sturgeon Acipenser brevirostrum gonad samples were collected from industry-reared fish and wild broodstock at various developmental stages to elucidate patterns of gonadal differentiation and maturation. Genital ridges, containing germ cells, were present in 26 day-old fish and distinct gonads were present by day 54. Sturgeon gonads are known to consist of two tissue types (adipose and gametogenic) and both were present at 72 day. Anatomical differentiation of gonads occurred by 6 months and was advanced by 15 months. Ovaries had distinct lamellae while testes remained non-lamellate. Gonial proliferation had occurred by 15 months, but the cells were not identifiable as spermatogonia or oogonia. Small white 'pinhead' oocytes were macroscopically visible in ovaries as early as 36 months. At 43 months ovaries were clearly organized, with some areas containing only immature oocytes and other containing oocytes apparently developing as cohorts. Individual fish showed considerable variation: the level of development remained unchanged at 84 months in some females, while others showed clear progression towards sexual maturation at 48 months. Sperm cells were present in males as early as 52 months. Advanced development of ovarian follicles was observed only in biopsies of re-conditioned broodstock of wild origin. In the year before spawning, the most advanced oocytes became pigmented, the chorion thickened, the nucleus (germinal vesicle) migrated towards the micropyle complex at the animal pole, and ovulation occurred in May under appropriate environmental conditions.     

Histological structure of the female gonads and ovipositor of the European bitterling,Rhodeus amarus (Bloch, 1782) (Cyprinidae: Acheilognathinae)

We investigated the histological structure of the female gonads and ovipositor of the European bitterling, Rhodeus amarus. The base of the ovipositor was formed by the conical organ or ‘eminence’. Based on the structure of the conical organ, including a special distribution of collagenous and muscular fibrils, a well‐developed vascular system and numerous scyphoid mucous cells in the internal epithelium, this organ was used for temporary storage of oocytes during the spawning period and for their movement along the ovipositor. An extensive network of blood vessels, muscular fibrils and numerous collagenous fibrils in the connective tissue of the ovipositor may make a functional contribution to the ovipositor by making it firmer during egg laying. Mucous cells were detected in the medial and distal regions of the ovipositor, which may play a role in facilitating insertion of the ovipositor into the exhalant siphon of a mussel during oviposition. European bitterling are batch spawners, and the female spawns eggs in clutches at intervals during the breeding season, which were visible as three distinct cohorts of oocytes in the ovary.