Gonadal sex differentiation in Atlantic halibut

The process of gonadal sex differentiation in 338 Atlantic halibut Hippoglossus hippoglossus larvae, ranging in size from 10 mm L_s to 230 mm L _F, is described histologically. Gonadal sex differentiation occurred by 38.0 mm L _F, which coincided with the weaned, post-metamorphic, settled stage in the life cycle. This was a gradual process that coincided with other organogenesis in the developing larvae.     

A first glimpse of larval ecology of halibut species in the Gulf of St. Lawrence,Canada

Knowledge of the larval ecology of winter-spawning fish from the Estuary and Gulf of St. Lawrence, Canada, remains scarce due to the seasonal ice cover that prevents ichthyoplankton sampling using conventional methods. Two winter-spawning species, Atlantic halibut (AH, Hippoglossus hippoglossus) and Greenland halibut (GH, Reinhardtius hippoglossoides), support the most important groundfish fisheries of this area. In March 2020, the authors captured 10 halibut larvae ranging in size from 5 to 14 mm during an opportunistic survey in the GSL onboard an icebreaking vessel. Of these, eight were AH and two GH. Judging by their very small size, the larvae were only a few days old, suggesting that the spawning grounds are close to the capture sites. This effort constitutes a first step in validating the putative spawning areas for these two important GSL stocks. This knowledge is important for the conservation and sustainable management of these fisheries.     

Astaxanthin binding to solubilized muscle proteins of Atlantic salmon (Salmo salar L.), haddock (Melanogrammus aeglefinus L.) and Atlantic halibut (Hippoglossus hippoglossus L.)

A study was conducted to compare astaxanthin binding ability of solubilized muscle proteins of Atlantic salmon (Salmo salar L.), haddock (Melanogrammus aeglefinus L.) and Atlantic halibut (Hippoglossus hippoglossus L.). Muscle proteins of juvenile Atlantic salmon, haddock and halibut were solubilized by sequential extraction of muscle tissue using low ionic strength solutions. Electrophoretic protein profiles of the six solubilized fractions from these species were similar. Each solubilized fraction from the three species was examined for its relative astaxanthin binding capacity. The amount of bound astaxanthin was significantly different (P < 0.05) among the six fractions of each species. Significant differences in astaxanthin binding were only found for fractions A and E among the species. The amount of bound astaxanthin in various fractions of each species showed a good correlation (R² = 0.80–0.92) with the ANS (8-anilino-1-naphthalenesulfonate) fluorescence intensity of those fractions. The pattern and extent of astaxanthin binding to the muscle proteins of juvenile salmon, haddock and halibut is comparable to that reported previously for adult Atlantic salmon [Saha, M.R., Ross, N.W., Gill, T.A., Olsen, R.E., Lall, S.P., 2005. Development of a method to assess binding of astaxanthin to Atlantic salmon S. salar L. muscle proteins. Aquacult. Res. 36, 336–343.]. These combined observations suggest that the carotenoid binding capacity of the muscle proteins of salmon is not the limiting factor in the deposition of carotenoid in their flesh.     

Gonadal sex differentiation and effect of rearing temperature on sex ratio in black rockfish (Sebastes schlegeli)

Temperature-dependent sex determination has been demonstrated in some species of fish, and a high temperature during the period of sex differentiation typically produces a male-dominant population. This research investigated the gonadal sex differentiation and effect of rearing temperature on the sex ratio in larval black rockfish Sebastes schlegeli, which is a viviparous species. Two types of gonads were histologically distinguishable in fish 20 mm in total length (TL). The putative ovary started forming an ovarian cavity, while the putative testis was not clearly differentiated until 51 mm TL. In a temperature-controlled experiment, the proportions of females were 45% at 10°C, 46% at 14°C, 50% at 18°C, 63% at 22°C, and 83% (significantly different from 1:1 sex ratio) at 24°C. These results suggest that morphological sex differentiation in black rockfish occurs at approximately 20 mm in TL, and it is possible that high temperatures (24°C) induce not a male- but a female-dominant population in this species.     

Regulation of growth in turbot (Scophthalmus maximus Rafinesque) and Atlantic halibut (Hippoglossus hippoglossus L.): aspects of environment × genotype interactions

This review is aimed atelucidating the mechanisms that regulate growthin cultured juvenile Atlantic halibut (Hippoglossus hippoglossus) and turbot (Scophthalmus maximus) by evaluating thesignificance of environmental factors(temperature, photoperiod) and the interactionsbetween them. In addition, we examine growthproperties in three populations of juvenilehalibut and turbot in order to elucidate anygeographical differences in growth and growthefficiency in these species. Both temperatureand photoperiod have a significant andpersistent effect on growth rate in bothspecies. Temperature tolerance, demonstrated bya wide temperature range supporting maximalgrowth, increased with size. Fish subjected tocontinuous light exhibited faster growth thanthose experiencing a natural photoperiod or aconstant short day. Moreover, when thephotoperiod increased naturally with day-lengthor when fish were abruptly switched from beingreared on short-day conditions to continuouslight, a subsequent increase in growth rate wasobserved. This growth enhancing effect ofextended photoperiods was more apparent in ashort time scale in Atlantic halibut than inturbot, but both species show significantlong-term effects of extended photoperiods.Enhanced growth in fish in continuous light waspartly explained by higher growth efficiency.In both species, there was a significantinteraction between temperature andphotoperiod, suggesting that thegrowth-enhancing effect of continuous light isrelatively stronger at lower temperatures.Growth rate in both species was alsosignificantly influenced by the interaction oftemperature and fish size, as the optimaltemperature for growth decreased rapidly withincreasing fish size. Differences in growth,food intake, food conversion efficiency,metabolism, ammonia excretion and RNA/DNAratios in white muscle were observed betweendifferent strains of halibut and turbot, withthe best growth properties being observed inthe northern populations. These findings onhalibut and turbot partly support the theory ofcountergradient variation in growth, suggestingthat populations from high latitudes havehigher growth capacity than populations fromlow latitudes.     

Genetic sex identification and the potential evolution of sex determination in Pacific halibut (Hippoglossus stenolepis)

The discovery of genetic markers linked to physiological traits in wild populations is increasingly desired for ecological and evolutionary studies, as well as to inform management decisions. However, identifying such markers often requires a large investment of both time and money. Serendipitously, in a recent microsatellite survey, we discovered three out of 16 microsatellite loci that were correlated to the female sex in Pacific halibut (Hippoglossus stenolepis). These three loci were screened in 550 Pacific halibut to determine their accuracy at identifying sex. Genetic assignment successfully identified sex in 92% of individuals from sample collections spanning 3,000 km and 9 years. All but two of 287 females had one copy of a characteristic allele for at least one of the three microsatellite loci, resulting in consistent heterozygote excess in females. This pattern is consistent with the hypothesis that females are the heterogametic sex in Pacific halibut, which thus may have a different sex-determination pattern than the closely related Atlantic halibut (Hippoglossus hippoglossus). A rapid divergence of sex-determining mechanisms could be either a cause or consequence of speciation between Pacific and Atlantic halibut. In either case, the ability to genetically identify sex in individual Pacific halibut provides a new tool for ecological studies, fisheries management, and insight into the evolution of sex determination in flatfish.     

Microsatellite analysis indicates an absence of population structure among Hippoglossus hippoglossus in the north‐west Atlantic

To explore the population structure of Atlantic halibut Hippoglossus hippoglossus , 160 fish from four locations in the north‐west Atlantic (Bay of Fundy, Scotian Shelf, Gulf of St Lawrence and Iceland) were examined for evidence of population structure using 18 microsatellite markers. Pair‐wise F _ST and a model‐based cluster analysis revealed no significant differentiation between samples, although uncertainties surrounding Atlantic halibut reproductive behaviour made it difficult to ascertain that only a single breeding population had been sampled at each location     

Microarray analysis of defined Mycobacterium tuberculosis populations using RNA amplification strategies

Background

Halibuts are commercially important flatfish species confined to the North Pacific and North Atlantic Oceans. We have determined the complete mitochondrial genome sequences of four specimens each of Atlantic halibut (Hippoglossus hippoglossus), Pacific halibut (Hippoglossus stenolepis) and Greenland halibut (Reinhardtius hippoglossoides), and assessed the nucleotide variability within and between species.

Results

About 100 variable positions were identified within the four specimens in each halibut species, with the control regions as the most variable parts of the genomes (10 times that of the mitochondrial ribosomal DNA). Due to tandem repeat arrays, the control regions have unusually large sizes compared to most vertebrate mtDNAs. The arrays are highly heteroplasmic in size and consist mainly of different variants of a 61-bp motif. Halibut mitochondrial genomes lacking arrays were also detected.

Conclusion

The complexity, distribution, and biological role of the heteroplasmic tandem repeat arrays in halibut mitochondrial control regions are discussed. We conclude that the most plausible explanation for array maintenance includes both the slipped-strand mispairing and DNA recombination mechanisms.     

Spatial distribution and life‐history aspects of blackspot seabream Pagellus bogaraveo (Osteichthyes: Sparidae)

Spatial distribution and life history aspects of Pagellus bogaraveo in the eastern Ionian Sea were investigated using the data from 13 different studies carried out in the area from 1983 to 2010. The spatial patterns of the abundance, biomass and mean size showed that the species inhabits the shallow waters of the shelf (<170 m depth) as juveniles up to a certain size (<180 mm total length, L_T), moving to deeper waters of the slope (mainly 400–500 m depth) as adults. The spatial pattern of abundance indicated a continuous distribution of the species in deep waters, with hot‐spot areas of high values, whereas in shallow waters distribution was more discontinuous, with higher concentrations of juveniles in estuaries and brackish waters. The study of biological aspects of the species revealed (1) a difference in the increase in mass between males and females, (2) protandrous hermaphroditism, accompanied by the presence of primary females and males that do not change sex, (3) a sex ratio in favour of females >250 mm L_T, (4) the presence of hermaphrodites between 200 and 370 mm, (5) a long reproduction period from June to March, (6) a size at first maturity around 300 mm and (7) a diet composition of adults based mainly on fishes, and also on opportunistic behaviour in the food scarce environment of deep waters. The results suggest that the species' distribution and feeding strategies are the most appropriate for the oligotrophic eastern Ionian waters and that these conditions result in smaller sizes of the species in the east Mediterranean Sea compared to the west basin and the east Atlantic Ocean, with implications for the growth and reproductive biology of the species.     

Observations on an Intranuclear Microsporidian in Lymphoblasts from Farmed Atlantic Halibut Larvae (Hippoglossus hippoglossus L.)

ABSTRACT. An intranuclear microsporidian was observed in lymphoblasts from the kidney of farmed Atlantic halibut larvae ( Hippoglossus hippoglossus ). In addition to spores, intranuclear pre-spore stages were observed. Uninucleated stages with a slight thickening of the cell membrane were the most simple developmental stages observed. Multinucleated developmental stages contain electron-lucent vesicles and electron-dense discs. The spores are ovoid and measure 2.9 × 1.2 μm (mean). Sporophorous vesicles, diplokarya and mitochondria were not observed in any of the developmental stages. The developmental stages observed are similar to those of the family Enterocytozoonidae, genus Enterocytozoon. However, there are several differences between the present species and E. bieneusi from enterocytes in man. The relationship to the other species in the genus, E. salmonis can only be determined when all the developmental stages of the species from Atlantic halibut are identified.     

Size at sex change and reproductive cycle of the limpets Patella vulgata and Patella ulyssiponensis (Mollusca: Patellogastropoda) from intertidal rocky shores of the Algarve coast (southern Portugal)

ABSTRACT

This study describes the reproductive cycle of the common limpet (Patella vulgata) and rough limpet (Patella ulyssiponensis) from the Algarve coast (southern Portugal). Individuals of both species were sampled monthly during two consecutive years, and subjected to gonad histology and calculation of the mean gonadal index (GI). Both species had balanced sex ratios (P. vulgata = 1M: 0.98F; P. ulyssiponensis = 1M: 1.03F), similar size-frequency distribution between sexes, and equivalent mean shell lengths (SL) and total weights (TW) between males and females. In these protandrous hermaphrodites, the estimated size at sex change was slightly smaller in P. vulgata (SL₅₀ ≈ 29 mm) than in P. ulyssiponensis (SL₅₀ ≈ 36 mm). The reproductive cycles of both species were characterised by a main spawning season in early spring to early summer, sometimes including subsidiary spawning events. A comparison of the main spawning season throughout the species distributional ranges revealed that populations of P. vulgata and P. ulyssiponensis from southern Portugal have later spawning periods than other populations from the northeastern Atlantic Ocean. The present information is relevant for conservation purposes and for the proposal of management measures for the harvesting activity targeting P. vulgata and P. ulyssiponensis in southern Portugal.     

Ten polymorphic microsatellite loci for the Atlantic halibut (Hippoglossus hippoglossus) and cross-species application in related species

In the present study, 10 polymorphic microsatellite DNA loci from Atlantic halibut (Hippoglossus hippoglossus) were isolated and characterized. The number of alleles for these loci ranged from 2 to 4 in tested 24 individuals. Observed and expected heterozygosities per locus varied from 0.21 to 0.70 and from 0.31 to 0.65, respectively. Most of these 10 microsatellite loci were successfully amplified and showed polymorphic in five related species. These loci will be useful for the assessment of genetic diversity and population structure of Atlantic halibut. H. Ding and C. Shao contributed equally to this work.     

Thyroxine as a Mediator of Metamorphosis of Atlantic Halibut, Hippoglossus Hippoglossus

The response of morphological, histological and endocrinological development to exogenous 1-thyroxine (T₄) and to water depth during metamorphosis in Atlantic halibut, Hippoglossus hippoglossus, was investigated. Exogenous T₄ was given in daily doses of 0.1, 0.05 ppm or a control treatment to halibut larvae at 550 daydegrees (posthatch, premetamorphic) for 14 days. Water depths of 40 cm, 10 cm or 1.5 cm were used to rear halibut larvae from 590 daydegrees for 21 days. Halibut larvae given exogenous T₄ at 0.1 ppm had accelerated eye migration relative to MH in fish given 0.05 ppm and in control fish. Pigmentation was correlated with dosage after 14 days. The volume of thyroid tissue was expressed in a dose-dependent manner and exhibited a size-dependency within each treatment. However, the follicles were atypical with reduced colloid, increased lumen and low epithelial cells even in the control group. The results indicate that T₄ is a mediator in halibut metamorphosis. In the water depth experiment, only cortisol levels of larvae reared in 1.5 cm water were significantly affected after 21 days, but this was not correlated with metamorphic rate. Hormone profiles, morphological changes and size suggest the existence of a window of opportunity for metamorphosis in halibut extending from about 16 mm and tapering off about 21 mm SL. The pooled hormone profiles indicate the commencement of a hormonal cascade similar to that of other flatfishes during metamorphosis. The results indicate that growth, neural and skeletal transformation, and pigmentation are biochemically separate processes in the metamorphosis of Atlantic halibut.     

Ichthyobodo hippoglossi n. sp. (Kinetoplastea: Prokinetoplastida: Ichthyobodonidae fam. nov.), an ectoparasitic flagellate infecting farmed Atlantic halibut Hippoglossus hippoglossus

Diseased Atlantic halibut Hippoglossus hippoglossus juveniles from a hatchery in western Norway showed gill and skin infections with an Ichthyobodo species. Genus Ichthyobodo contains a single valid species, I. necator, a parasite originally described from the skin and fins of a salmonid fish in freshwater. Many studies have identified this species from other hosts, but recent molecular evidence suggests that many Ichthyobodo spp. occur in both fresh- and seawater. We redescribe I. necator from Atlantic salmon Salmo salar skin infections in Norway and compare the morphology of I. necator with the form from halibut. A scheme to standardise the measurements of Ichthyobodo cells is presented. Morphologically, the Ichthyobodo species from the skin and gills of halibut differs from I. necator from salmon skin by shape (in air dried stained smears), by a low number of variably sized kinetoplasts and by a long flagellar pocket. There is also a clear increase in the number of kinetoplasts in L necator with increasing cell size (area), a pattern absent from Ichthyobodo sp. from halibut. The 2 forms are also clearly separated by their small subunit (ssu) rDNA sequences; alignments of partial ssu sequences showed 93.5 % similarity. Consequently, Ichthyobodo sp. from halibut is considered a new species, and is named I. hippoglossi n. sp. Its closest relative is Ichthyobodo sp. IV from another marine fish, the Atlantic cod Gadus morhua. A family, Ichthyobodonidae fam. nov. in the order Prokinetoplastida Vickerman, 2004, is erected to encompass Ichthyobodo spp.     

Vulnerability to longline fisheries of three hammerhead shark Sphyrna species in the south‐western and equatorial Atlantic Ocean

Catch and effort data from 29 418 longline sets from Brazilian tuna longline vessels operating in the south‐western and equatorial Atlantic Ocean between 2004 and 2011 were analysed to investigate the distribution, catch rate and size of three species of hammerhead sharks (Sphyrna lewini, Sphyrna mokarran and Sphyrna zygaena). During that period, 6172 hammerhead sharks were caught. Among the elasmobranchs, the highest percentage of hammerhead sharks were caught in 2007, when they accounted for 3·90% of the group, while the lowest value of 0·40% was recorded in 2010. In general, the spatial distribution of the mean catch per unit effort (CPUE) by years and quarters showed a trend of higher catches near the equatorial region and in southern Brazil. The nominal mean CPUE was 0·12 Sphyrna spp. 1000⁻¹ hooks, with the highest value being recorded in 2007 (0·30 Sphyrna spp. 1000⁻¹ hooks). The standardized yearly CPUE estimated by a generalized linear model assuming a zero inflated negative binomial (ZINB) distribution were not much different from nominal values. Of the 205 sexed specimens, 117 were females and 88 were males, resulting in a sex ratio with a predominance of females (1·30:1·00), although not statistically significant. The total length of females ranged from 1200 to 2800 mm and of males from 1100 to 3100 mm. Juvenile hammerhead sharks represented 82 and 54% of the sexed female and male specimens, respectively.     

Real-time PCR analysis of ovary- and brain-type aromatase gene expression during Atlantic halibut (Hippoglossus hippoglossus) development

Two forms of cytochrome P450 aromatase, acting in both the brain and the ovary, have been implicated in controlling ovarian development in fish. To better understand the expression of these two enzymes during sexual differentiation in Atlantic halibut (Hippoglossus hippoglossus), real-time PCR was used to quantify the mRNA levels of ovary- (cyp19a) and brain-type cytochrome P450 aromatase (cyp19b) genes in the gonad and brain during gonadal development. Both enzymes showed high levels of expression in both tissues in developmental stages prior to histologically detectable ovarian differentiation (38 mm fork length), with increased expression occurring slightly earlier in the brain than the gonad. Cyp19a showed a second peak of expression in later stages (> 48 mm) in the gonad, but not the brain. Cyp19b expression was generally higher in the brain than the gonad. These results suggest that sexual differentiation may begin in the brain prior to gonadal differentiation, supporting the idea that steroid hormone expression in the brain is a key determinant of phenotypic sex in fish. In an examination of sexually immature adults, cyp19a was highly expressed in female gonad while cyp19b was very highly expressed in the pituitary of both sexes. The ratio of cyp19a to cyp19b expression was much higher in ovaries than in testes in the adult fish, so this ratio was analyzed in the developing gonads of juvenile halibut in an attempt to infer their sex. This was only partially successful, with about half the fish in later developmental stages showing apparently sex-specific differences in aromatase expression.     

Temperature effects on sex differentiation of two South American atherinids, Odontesthes argentinensis and Patagonina hatcheri

Synopsis The present study investigated the effects of water temperature (18, 21, and 25 °C) on the histological process of gonadal sex differentiation of two commercially important atherinid fishes from South America, Odontesthes argentinensis (sea pejerrey) and Patagonina hatcheri (Patagonian freshwater pejerrey). In both species, female gonadal sex differentiation began with the formation of lateral stromal cell outgrowths and the appearance of meiotic oocytes. The male gonads remained quiescent for about twice as long as the female gonads, with differentiation becoming evident by the formation of the main sperm duct and of cysts of germ cells at the periphery of the gonads. Meiosis in males occurred relatively long after somatic differentiation of the testis. The ovaries of O. argentinensis differentiated at 28 days (20.3 mm) at 25 °C, 42 days (24.0 mm) at 21 °C, and 56 days (23.8 mm) at 18 °C. In the males, differentiation was observed at 98 days at 25 and 21 °C (39.4 mm and 40.4 mm, respectively), but at 112 days under 18 °C (40.7 mm). In P. hatcheri, differentiation of females occurred at 21 days (17.8 mm) at 25 °C, 28 days (20.8 mm) at 21 °C, and 35 days (23.2 mm) at 18 °C. Male differentiation became evident at 56 days under 25 and 21 °C (30.8 and 32.7 mm, respectively), and at 70 days (37.7 mm) at 18 °C. The sex-ratios of O. argentinensis reared at 18 or 21 °C were female-biased whereas those at 25 °C were not; groups reared at 18 °C had significantly more females than groups from the same progeny reared at 25 °C. In contrast, the sex-ratios in all groups of P. hatcheri did not differ significantly from 1:1 and no significant differences were found between groups of the same progeny reared at different temperatures. These results suggest the occurrence of thermolabile sex determination (TSD) in O. argentinensis whereas in P. hatcheri gonadal sex appears to be strongly genetically determined.