Seasonal patterns of winter flounder Pseudopleuronectes americanus abundance and reproductive condition on the New York Bight continental shelf

To resolve varied and sometimes conflicting accounts of spawning and habitat characteristics for winter flounder Pseudopleuronectes americanus, seasonal patterns in abundance and reproductive condition were investigated in the New York Bight, near the southern edge of their current reproductive range. Fish were collected from trawl surveys on the inner continental shelf from October 2006 to October 2007. Pseudopleuronectes americanus were most abundant during January and April surveys, were rarely collected in August, with intermediate abundances in June and October. Measurements of fish condition [hepato‐somatic index (I_H), condition factor (K) and the per cent dry mass of muscle tissue (%M_D)] and reproductive condition [gonado‐somatic index (I_G)] were determined to evaluate seasonal changes in energy accumulation and depletion and reproduction. Males and females had similar patterns in body and reproductive condition, although the magnitude of change was greater for females. I_H values were highest during spring and early summer, suggesting increased feeding following spawning. K and %M_D increased through spring and summer then declined in the autumn and winter concurrent with gonadal development. Gonads began developing in the autumn, and in January, I_G values approached spawning levels, with many spent individuals collected in spring. Within these general patterns, however, there was a large degree of variability among individuals, and a few mature non‐reproductive (‘skipped spawning’) females were observed. In the period after spawning, increased energy intake, indicated by increased I_H, may influence reproductive output since this energy is gradually transferred to the muscle and used for gonadal development in the forthcoming year. The occurrence of ripening individuals on the inner continental shelf in January suggests that these fish either rapidly move into estuaries to spawn by February–March or they remain on the inner shelf to spawn, or some combination of these. Future studies should evaluate these possibilities, as both estuarine and inner shelf habitats are potentially affected by activities such as dredging, sand dredging and wind energy development.     

Older triploid fish retain impaired reproductive endocrinology in the European sea bass Dicentrarchus labrax

This paper reports on an evaluation of growth, gonadal development and reproductive endocrinology of older triploid (3n) European sea bass Dicentrarchus labrax in comparison with their diploid (2n) counterparts throughout their fifth and seventh annual cycle of life. While older triploids retained impaired reproductive endocrinology, a sexually related dimorphic growth was observed with 3n females attaining the largest sizes. Comparisons of some body indexes showed that 3n females had a significantly lower hepato‐somatic index (I_H) than 2n females but a significantly higher viscero‐somatic index (I_F). In contrast, both male and female triploids showed significantly lower gonado‐somatic index (I_G) than diploids. Accordingly, diploids produced mature gametes but triploids did not, demonstrating that despite the longer time given to triploids for gonadal development, they could not reproduce. Furthermore, older triploids had lower levels of plasma sex steroids (testosterone, T; 11‐ketotestosterone, 11‐KT and oestradiol‐17β, E₂) and luteinizing hormone (LH) than their 2n counterparts with 3n females showing drastic effects of triploidization on their reproductive endocrinology. Vitellogenin (VTG) was undetectable in 3n females. Gonadal content of steroid hormones and Sparus aurata‐type gonadotropin‐releasing hormone (sbGnRH) in the brain and pituitary were also lower in triploids compared with diploids. These results suggest that older 3n D. labrax retain functional sterility in both sexes, and 3n females might reach larger sizes than 3n males and their 2n counterparts in this species.     

Comparative study of reproductive biology in single and multiple-spawner cyprinid fish. II. Sex steroid and plasma protein phosphorus concentrations

The dynamics and regulation of oogenesis in single and multiple-spawner cyprinid fish showing group-synchronous oocyte development, was investigated in three species from the River Meuse (Belgium): the roach Rutilus rutilus as a single spawner, and the bleak Alburnus alburnus and the white bream Blicca bjoerkna as multiple spawners. This paper compares the seasonal profiles of sex steroids (oestradiol-17β, testosterone and 17,20β-dihydroxy-4-pregnen-3-one) and plasma alkali-labile protein phosphorus. Different patterns of plasma oestradiol-17β (E2) and plasma protein phosphorus (PPP) have been observed not only between the single and the multiple spawner fish, but also among the two multiple spawners. In roach, two increases of E2 levels were observed. The first occurred in September after a short gonadal quiescent period, and coincided with the increase of the PPP at the onset of exogenous vitellogenesis. The second took place in spring before the spawning season. The low PPP recorded during that period probably reflected its rapid incorporation by the oocytes. In both multiple spawners, highest values of PPP were recorded just before the spawning season. In white bream, the PPP declined progressively once the differentiation of exogenous vitellogenic oocytes was completed before the onset of the spawning season. In bleak, PPP levels remained high throughout the spawning season and corresponded to a sustained oocyte recruitment during the whole of this period. Regardless of the pattern of oocyte growth recruitment, the E2 concentrations were high in both multiple spawner species during the breeding season. In the three species, testosterone levels remained low regardless of the maturation stage (ranging from 0.6 to 13.4ng ml^?1). Except for relatively high concentrations of 17,20βP in roach during final maturation and postspawning stages (20 and 28 ng ml^?1, respectively), low levels of this steroid were measured in these cyprinids, and especially in the multiple spawner fish. The role of this progestogen as the maturation inducing Steroid is discussed.     

The reproductive cycle of female Ballan wrasse Labrus bergylta in high latitude,temperate waters

This 2 year study examined the reproductive cycle of wild female Ballan wrasse Labrus bergylta in western Norway as a precursor to captive breeding trials. Light microscopy of ovarian histology was used to stage gonad maturity and enzyme‐linked immuno‐absorbent assay (ELISA) to measure plasma concentrations of the sex steroids testosterone (T) and 17β‐oestradiol (E₂). Ovarian recrudescence began in late autumn to early winter with the growth of previtellogenic oocytes and the formation of cortical alveoli. Vitellogenic oocytes developed from January to June and ovaries containing postovulatory follicles (POF) were present between May and June. These POF occurred simultaneously among other late maturity stage oocytes. Plasma steroid concentration and organo‐somatic indices increased over winter and spring. Maximal (mean ±s.e .) values of plasma T (0·95 ± 0·26 ng ml^?1), E₂ (1·75 ± 0·43 ng ml^?1) and gonado‐somatic index (I_G; 10·71 ± 0·81) occurred in April and May and decreased greatly in July when only postspawned fish with atretic ovaries occurred. Evidence indicates that L. bergylta are group‐synchronous multiple spawners with spawning occurring in spring and peaking in May. A short resting period may occur between late summer and autumn when previtellogenic oocytes predominate and steroid levels are minimal.     

Reproductive biology of hardhead catfish Ariopsis felis: evidence for overwintering oocytes

Hardhead catfish Ariopsis felis are a common marine catfish in the coastal waters of the Gulf of Mexico (GOM). The low economic value of this species has depressed interest and research, and although the species is known for its extremely low fecundity and large oocytes, little else is known about this catfish species. A total of 1230 samples across all months of the year from 2016 to 2018 resulted in 681 females, and analysis of gonado-somatic index (I_G) revealed 1% to be a clear cut-off indicating maturity. Females are considered capable of spawning from April to June when I_G averaged 4–8%. Both atresia and post-ovulatory follicles were present in July, suggesting that spawning ends in July in the northern GOM. The 1% I_G cut-off was used to designate maturity, and from that an L₅₀ of 253 mm was estimated. Batch fecundity from 41 females estimated a mean batch size of 36 oocytes. Perhaps the most interesting finding was the presence of secondary growth stage oocytes (e.g., cortical alveoli) from July through November, well outside the spawning capable period. Furthermore, 78% of females had some early vitellogenic oocytes present during the non-spawning season, and the distribution of these relatively large (2–5 mm) oocytes did not change over time. The results here are not only important as reproductive biology information for a common and abundant species, but also present interesting and unusual patterns of non-spawning season oocyte development that is not commonly seen in Western Hemisphere subtropical fish species.     

Spatial and temporal variation in predation on reef fishes by coral trout (Plectropomus leopardus,Serranidae)

The diet of coral trout Plectropomus leopardus (Serranidae) was studied over a two year period at One Tree Island, Great Barrier Reef, Australia. Rapid visual counts demonstrated that P. leopardus were most abundant on the reef slope habitat and inner edge of the enclosed lagoon. Few P. leopardus were found at sites from inner lagoon. It was hypothesized that diet would vary among habitats, times and size classes of coral trout. Ninety-two percent of P. leopardus that contained prey had consumed fish and 87% had only eaten fish. Many types of reef fish were taken by P. leopardus (e.g. Pomacentridae, Scaridae, Blenniidae and Labridae). Most pelagic prey (Clupeidae and Engraulididae) were taken on the reef slope, while some prey were solely or pimarily taken in the lagoon (e.g. Blenniidae and crustaceans). Most pelagic prey were taken on the reef slope in summer by P. leopardus>250 mm (SL). Plectropomus leopardus (<200 mm) from the lagoon had a higher proportion of invertebrates in the diet than fish from the reef slope. Plectropomus leopardus of all sizes ate small fish, while largest fish generally consumed largest prey (especially adult scarids and labrids). I argue that interactions among multiple species of prey and predators need more attention, because piscivores may respond to prey in different ways according to habitat type as well as the number and type of other prey types present. Furthermore, different sizes of fish (e.g. coral trout) may impact assemblages of prey in different ways.     

Seasonal changes in adrenal and gonadal activity in the quail, Perdicula asiatica: involvement of the pineal gland

The present study assessed annual adrenal gland activity in the Indian tropical Jungle bush quail, Perdicula asiatica. We also elucidated the role of the annual variations in gonadal steroids and melatonin in the regulation of its activity. Increasing day length (photoperiod), ambient temperature and rainfall are positively correlated with adrenal and gonadal functions, and inversely related to pineal gland activity. Pineal, adrenal and gonadal weights showed cyclical patterns relative to environmental factors, which were also correlated with plasma melatonin, corticosterone and gonadal steroids, respectively. In both sexes of P. asiatica, pineal gland weight and/or plasma melatonin levels were inversely related to adrenal lipids, (e.g. phospholipids, free and esterified cholesterol) and plasma corticosterone levels. Melatonin levels also showed an inverse relationship with plasma testosterone and estradiol levels. These studies indicate that changes in environmental factors promote annual variations in adrenal and gonadal activity probably by modulating the pineal gland. Melatonin receptors have been localized in the pars tuberalis, adrenal gland and gonads of birds, the pineal gland may, therefore, mediate environmental stimuli indirectly and directly to down regulate adrenal and gonadal activity, which run in parallel in this species.     

De novo sequencing and chromosomal‐scale genome assembly of leopard coral grouper,Plectropomus leopardus

The leopard coral grouper, Plectropomus leopardus, belonging to the family Epinephelinae, is a carnivorous coral reef fish widely distributed in tropical and subtropical waters of the Indo‐Pacific. Due to its appealing body appearance and delicious taste, P. leopardus has become a popular commercial fish for aquaculture in many countries. However, the lack of genomic and molecular resources for P. leopardus has hindered study of its biology and genomic breeding programmes. Here we report the de novo sequencing and assembly of the P. leopardus genome using a combination of 10 × Genomics, high‐throughput chromosome conformation capture (Hi‐C) and PacBio long‐read sequencing technologies. The genome assembly has a total length of 881.55 Mb with a scaffold N50 of 34.15 Mb, consisting of 24 pseudochromosome scaffolds. busco analysis showed that 97.2% of the conserved single‐copy genes were retrieved, indicating the assembly was almost entire. We predicted 25,248 protein‐coding genes, among which 96.5% were functionally annotated. Comparative genomic analyses revealed that gene family expansions in P. leopardus were associated with immune‐related pathways. In addition, we identified 5,178,453 single nucleotide polymorphisms based on genome resequencing of 54 individuals. The P. leopardus genome and genomic variation data provide valuable genomic resources for studies of its genetics, evolution and biology. In particular, it is expected to benefit the development of genomic breeding programmes in the farming industry.     

Reproductive effort,fecundity and energy allocation in two species of the genus Perinereis (Polychaeta: Nereididae)

Summary

The reproductive effort in terms of fecundity and energy allocation was studied in two species of semelparous polychaetes belonging to the genus Perinereis, living in the same environment, with different reproductive modalities. There is a great individual variability both in terms of reproductive effort and fecundity. Fecundity varied from 4080 to 15000 oocytes in P. rullieri and from 7000 to 26000 in P. cultrifera; no linear relationship was found between oocyte number and total jaw length utilised as size index. The energy content of germinal and somatic tissues was determined by Differential Scanning Calorimeter (DSC). The reproductive effort was calculated as RE = E_G/(E_G + E_S) where E_G is the total energy in germinal tissues and E_S is the total energy in somatic tissues. Reproductive effort is very high with mean values of 0.62 for P. rullieri and 0.79 for P. cultrifera. The different amounts of energy allocated in germinal tissues can be attributed to the different reproductive modalities—P. rullieri reproduces in the atokous phase whereas P. cultrifera has conserved epitoky in its life-cycle. The lack of correlation between reproductive effort and size index strongly suggests that reproductive allocation does not increase with age. In semelparous species the variability in fecundity and reproductive effort observed cannot be interpreted in terms of a trade-off between fecundity and survival as in iteroparous species. In fact, in semelparous an individual allocates all available resources to reproduction and then dies.     

Reproduction in puyen, Galaxias maculatus (Pisces: Galaxiidae), in the southernmost extreme of distribution

Reproductive aspects of a peripheral population of Galaxias maculatus are described and the relationship with the physiology and handling of energetic reserves under marginal environmental conditions is investigated. The G. maculatus population of Tierra del Fuego has an extended reproductive season, with differences in timing and duration compared to other populations of continental Patagonia, New Zealand and Australia. Elevated gonadosomatic indexes (I_G) were observed during this period in both sexes (maximum I_G = 33.49% males; 35.94% females). The high abundance of mature males (with high I_G values) on the spawning grounds during the reproductive season suggests that they were waiting for the return of the mature females. Larger females reached total maturation at the beginning of the reproductive season, whereas the size of maturing females diminished toward the end (mean TL = 96 mm, October; 70 mm, February). Both sexes showed an extremely high investment in reproduction, reaching a maximum I_G of about 35% for both sexes. At the beginning of the reproductive season females reached the maximum median I_H (3.37%) and males the minimum (0.96%), suggesting differences in the role of the liver in the management of energetic reserves during sexual maturation. The variation in the fat index (I_F) suggests that fat reserves were used to survive winter (maximum median I_F > 1%, autumn; minimum about 0.2%, spring).     

Reproductive cycles in a reared strain of the mummichog, a daily spawner

Annual, lunar, and diel samplings were taken from a strain of mummichog (Arasaki strain) reared in outdoor tanks under natural conditions, to examine gonadal maturity. Gonads of yearling fish were quite immature in September. During late autumn and winter, a gradual increase in the GSI of both sexes was observed, and the growth of cortical alveolus phase oocytes in females and basal spermatogenesis in males progressed. In late February, a rapid increase in the GSI of both sexes, vitellogenesis in females, and active spermatogenesis in males, occurred. The spawning period of the yearling fish was from late March to August judging from the presence of milt-producing males and ovulated females. The spawning period of the underyearling fish started in the same month as the yearlings, but terminated 1 month earlier. Plasma levels of oestradiol-17 β (E₂) in females and testosterone in males were high during the spawning period in the yearlings. In the underyearlings, however, E₂ levels peaked in early spring, and declined in the latter part of the spawning period. Neither a lunar nor semilunar cycle was evident in the reproductive activity of this fish, which proved to be a typical daily spawner. Females showed an apparent daily reproductive cycle; oocyte maturation commenced at about 1200 hours, germinal vesicle breakdown (GVBD) occurred at about 2400 hours, and ovulation was completed by 2400 hours, 24 h after GVBD. Such clear annual and daily reproductive cycles make this strain of mummichog a suitable model for the study of environmental and endocrine regulation of reproductive cycles in marine and estuarine teleosts.     

The adaptive potential of a plant pathogenic fungus, <Emphasis Type="Italic">Rhizoctonia solani</Emphasis> AG-3, under heat and fungicide stress

The ability to improve fitness via adaptive evolution may be affected by environmental change. We tested this hypothesis in an in vitro experiment with the plant pathogen Rhizoctonia solani Anastomosis Group 3 (AG-3), assessing genetic and environmental variances under two temperatures (optimal and higher than optimal) and three fungicide concentrations (no fungicide, low and high concentration of a copper-based fungicide). We measured the mean daily growth rate, the coefficient of variation for genotypic (I _G) and environmental variance (I _E) in growth, and broad-sense heritability in growth. Both higher temperature and increased fungicide concentration caused a decline in growth, confirming their potential as stressors for the pathogen. All types of standardized variances in growth—I _G, phenotypic variance, and I _E as a trend—increased with elevated stress. However, heritability was not significantly higher under enhanced stress because the increase in I _G was counterbalanced by somewhat increased I _E. The results illustrate that predictions for adaptation under environmental stress may depend on the type of short-term evolvability measure. Because mycelial growth is linked to fitness, I _G reflects short-term evolvability better than heritability, and it indicates that the evolutionary potential of R. solani is positively affected by stress.     

Reproductive effort,fecundity and energy allocation in Marphysa sanguinea (Polychaeta: Eunicidae)

Summary

Reproductive effort in terms of fecundity and energy allocation was studied in the iteroparous and long lived polychaete Marphysa sanguinea. Both measures show great variability. Fecundity varied from 8500 to 24300 oocytes; no linear relationship was found between oocyte number and jaw length whereas a direct relationship was established between oocyte number and wet body weight. The energy content of germinal and somatic tissues was determined by differential scanning calorimeter (DSC). The reproductive effort for a single reproductive event was calculated according to the formula: RE = E_G/(E_G + E_S) where E_Gis the total energy of the germinal tissues and E_S is the total energy of the somatic tissues. The lack of correlation between reproductive effort and size index strongly suggests that reproductive allocation does not increase with age. The reproductive effort ranged from 0.04 to 0.19 with a mean value of 0.120.     

Individual, temporal, and population-level variations in circulating 11-ketotestosterone and 17β-estradiol concentrations in the oyster toadfish Opsanus tau

Sex steroid hormones are important for reproduction in all vertebrates, but few studies examine inter-individual, temporal, and population-level variations, as well as environmental influences on circulating steroid levels within the same species. In this study we analyzed plasma 11-ketotoestosterone (11-KT) and 17β-estradiol (E₂) levels in the oyster toadfish to test for 1) individual and temporal variations by serially sampling the same individuals during the reproductive and post-reproductive period, 2) variations in steroid levels among toadfish obtained from different sources or maintained under different holding conditions, and 3) correlations with environmental parameters. Results from serial sampling showed marked inter-individual variations in male 11-KT levels in two separate groups of toadfish, but no temporal differences from June to September. Females also showed inter-individual variations in E₂ concentrations, but most had elevated levels late in the reproductive season coincident with oocyte growth prior to winter quiescence. E₂ concentration, but not 11-KT, was positively correlated with water temperature, and negatively correlated with daylength and lunar phase. Maricultured toadfish held under constant conditions had elevated levels of E₂ and 11-KT that should be considered when using these fish for experimentation. This study provides important comparative information on the relationship between individual variations in steroid levels, and how they relate to physiological and environmental correlates in a model marine teleost.     

Effects of exogenous estradiol-17β on early growth and gonadal development of diploid and triploid female rainbow trout (Oncorhyncus mykiss)

Triploidy is a viable condition in teleosts. However, in many salmonids, the triploid condition in the female results in sterility as gametogenesis appears to be disrupted. Although the underlying mechanisms regulating the gonadal development of teleosts have not been clearly elucidated, the reversal of phenotypic sex by the administration of the appropriate exogenous steroid during early development supports the argument that gonadal steroids play a pivotal role in sexual differentiation and subsequent gonad development in these fish. To determine whether the failure of normal ovarian development in triploid female rainbow trout (Oncorhynchus mykiss) is due to an absence or reduction of endogenous sex steroids, ovarian morphology was compared between diploid and triploid juvenile rainbow trout treated with exogenous estradiol-17β (E₂). The ovaries of both untreated and E₂ treated diploid fish, at 145 days post-fertilization, contained synchronously developing oocytes in the perinucleolar stage, whereas ovaries from untreated and estradiol-treated triploid fish of the same age were considerably smaller and devoid of developing oocytes. No differences in the ovaries of triploid untreated fish and triploid fish treated with E₂ were observed. It is reported that exposure to exogenous E₂ during the period of gonadal differentiation is not sufficient to induce oocyte development in triploid rainbow trout. © 1996 Wiley-Liss, Inc.     

Effect of fish length and nutritional condition on the fecundity of distressed Atlantic cod Gadus morhua from the Baltic Sea

The disappearance of larger individuals and the decrease in individual body condition suffered by Atlantic cod Gadus morhua in the eastern Baltic during the past two decades can be expected to affect the stock reproductive output. To investigate this, female G. morhua were collected during the spawning and pre‐spawning period in 2015?2016. The current individual potential fecundity (F_P) of eastern Baltic G. morhua was estimated and analysed in relation to total length (L_T) and indices of nutritional status such as body condition (K) and hepato‐somatic index (I_H) using generalized linear models. In addition, the current prevalence of atresia and its potential relation to K were investigated. Moreover, a calibration curve to estimate F_P from oocyte diameter, based on the autodiametric oocyte counting method, was established for the first time for eastern Baltic G. morhua and can be used for future fecundity studies on this stock. The results showed that F_P was mainly positively related to fish length, but K and I_H also contributed significantly to the variation in F_P. The model predicted that fish with K = 1·2 have a F_P 51% higher than fish of the same L_T with K = 0·8. The prevalence of fecundity regulation by atresia was 5·8%, but it was found only in fish in the pre‐spawning maturity stage and with low K. Temporal changes in biological features such as the length composition and individual body condition of eastern Baltic G. morhua, should be accounted for when estimating stock reproductive potential.     

Endocrine changes during the previtellogenic stage of the great sturgeon,Huso huso (Linnaeus, 1758)

The study aimed to investigate changes in serum concentrations of sex steroids, thyroid hormones, stress indices, cholesterol, triacylglycerol, calcium and phosphorus, as well as the growth in the 3‐year old previtellogenic stage of great sturgeon, Huso huso over a 6‐month period. Forty individually marked fish (mean weight 6913 g) were divided among three circular tanks under a natural temperature and photoperiod. Every 1.5 months fish were weighed and blood samples taken for serum analysis. Fish showed significant changes in 17β estradiol, testosterone, triiodothyronine (T₃), thyroxine (T₄), adrenocorticotropic hormone, cortisol, cholesterol, and phosphorus, but not in triacylglycerol and calcium concentrations. From the January onset of the experiment to the end of the trial in July, there was a decrease in sex steroids and cortisol concentrations as well as a decline in T₃ concentrations from March and in T₄ from February to July. The mean weight of fish increased significantly from the beginning to the end of the experiment. Although the oocyte diameter increased from 197 to 202 μm, the difference was not significant. The histological data on oocytes suggest that the great sturgeon, like the other sturgeons, has a heterogeneous asynchronous ovarian development in the early stage of reproduction. This data might be useful to the further understanding of the physiology of Huso huso and/or to improve its culture. Moreover, it leads to a general increase in the understanding of the basic reproductive biology of this valuable chondrostean species.     

Patterns of oocyte growth, vitellogenin and gonadal steroid concentrations in greenback flounder

The pattern of oocyte development in association with changes of plasma concentrations of vitellogenin (Vtg), 17β‐oestradiol (E₂) and testosterone (T) was investigated in maturing female greenback flounder Rhombosolea tapirina over the first part of a reproductive season (February to June). Examination of oocyte size‐frequency distributions showed that the oocyte developmental pattern in R. tapirina is multiple group synchrony, and that reproductively mature fish were present at all sampling times. There were no significant temporal variations in the gonado‐somatic index ( I _G), hepato‐somatic index ( I _H), or plasma concentrations of Vtg, E₂ and T during the sampling period, which indicates that reproductive development is not synchronized within the population. Significant increases in I _G, I _H and plasma concentrations of Vtg, E₂ and T, however, were observed in vitellogenic fish, and in fish undergoing final maturation. A positive relationship was also found between the growth of oocytes and plasma concentrations of Vtg, E₂ and T, although the patterns of increase were different for each variable. Plasma concentrations of Vtg and E₂ rose steadily across oocyte sizes from 100 to 450 μm, but the rate of increase of plasma E₂ was slower than that of Vtg, and both reached a saturated concentration at oocyte sizes of c . 450 μm. In contrast, plasma concentrations of T showed no marked increase until oocytes grew beyond 400 μm.     

Milt characteristics and plasma levels of gonadal steroids in greenback flounder Rhombosolea tapirina following treatment with exogenous hormones

Fish were treated with exogenous hormones, and milt and blood samples were collected for up to 96 h post‐treatment. Blood plasma samples were assayed for the gonadal steroids testosterone (T), 11‐ketotestosterone (11KT) and 17,20ß‐dihydroxy‐4‐pregnen‐3‐one (17.20ßP). Milt volume, spermatocrit and sperm motility were measured from milt samples. Non‐spermiated fish showed increased plasma T and 11KT in response to human chorionic gonadotropin (hCG) but not luteinising hormone releasing hormone analogue (LHRHa). Fish did not become spermiated in response to treatment with hCG, LHRHa, 11KT, 17‐hydroxyprogesterone (17P) or 17,20ßP. Spermiated fish showed an increase in milt volume in response to hCG and LHRHa but not exogenous steroids. Sperm motility declined to zero over 120 s and was not affected by hormone treatment or sampling time. Increased milt volume was accompanied by increased plasma T and 11KT, but not 17.20ßP levels. In a separate experiment, LHRHa delivered by injection or pellet was equally effective at increasing milt volume but had no effect on plasma steroid levels. Spermatocrit declined with stripping but was not affected by hormone treatment, nor was sperm motility. Co‐treatment of fish with 17P plus LHRHa had no additive effect on plasma steroid concentrations or milt volume. The results suggest that as in other teleosts, gonadotropin mediates milt production in greenback flounder.