Androgen secretion activity of recombinant follicle-stimulating hormone of Japanese eel, Anguilla japonica in immature and maturing eel testes

The androgen secretion activities of recombinant Japanese eel follicle-stimulating hormone (rjeFSH) were investigated in immature and maturing eel testes. The rjeFSH stimulated testosterone (T) and 11-ketotestosterone (11-KT) secretion in immature testis but not in maturing testis. This result suggests that eel FSH plays an important role through the sex steroid secretion in immature testis rather than in maturing testis.     

Exotic silver eels Anguilla anguilla in Japanese waters: seaward migration and environmental factors

To understand the ecology of the exotic silver European eel Anguilla anguilla introduced into Japanese waters, the migratory behavior of 106 specimens captured in the coastal waters of Japan between April 1997 and March 2002 was analyzed. Their migratory behavior was apparently correlated with various environmental factors, particularly photoperiod, water temperature, lunar phase, and passage of atmospheric depressions, and was similar to the behavior of the species in European waters. These findings suggest that transplanted European eels retain their ability to respond to environmental cues for seaward migration in similar temperate habitats. The timing of the migration of silver European eels coincided with that of the native Japanese eels A. japonica, suggesting that the silver European eel was synchronized physiologically with the native eel by the same environmental factors.     

鲤脑垂体匀浆液和人绒毛膜促性腺激_省略_性腺激素及性类固醇激素含量的影响

雄鳗注射５－６次、雌鳗注射９－１０次鲤脑体匀浆液（ＣＰＥ）＋人绒毛膜促性腺激素（ＨＣＧ）能分别诱导精巢和卵巢发育成熟。在雌雄鳗鲡,注射ＣＰＥ＋ＨＣＧ可显著增加端脑、间脑、中脑和下丘脑ｍＧｎＲＨ的含量,而对后脑和延髓ｍＧｎＲＨ的影响较小;注射ＣＰＥ＋ＨＣＧ增加雄鳗后脑和延髓ｃＧｎＲＨ－Ⅱ含量,对雌鳗脑区ｃＧｎＲＨ－Ⅱ则无显著影响。雌雄鳗鲡每次注射ＣＰＥ＋ＨＣＧ后１天,血清促性腺激素（ＧｔＨ）急剧上升     

Induction of spermatogenesis in Japanese eel by recombinant goldfish gonadotropins

We previously demonstrated the biological activities of single-chain recombinant gonadotropins (scGTHs) of goldfish Carrassius auratus follicle-stimulating hormone (scFSH) and luteinizing hormone (scLH), produced by a baculovirus–silkworm larvae system, by using in vivo bioassays with some fishes including Japanese eel Anguilla japonica. Among the bioassays, we succeeded in induction of spermatogenesis of sexually immature male Japanese eels by both scFSH and scLH, especially resulting in the occurrence of spermatozoa in scLH-administered males. However, those recombinant hormones did not induce enlargement of testes. In order to further confirm the potency of recombinant GTHs for use in aquaculture species, we administered scFSH and scLH to males of Japanese eel at higher dosage and frequency (eight times with 2–5 days interval) than those of the previous study (five or six times with 7 days intervals), including combination of scFSH and scLH administration (scFSH–scLH). Gonadosomatic indices (GSI) of scLH- and scFSH–scLH-administered males were larger than those of initial control males and of control males that were injected with saline. Enlargement of testes was also confirmed by measurement of testicular lobe size in scFSH-, scLH-, and scFSH–scLH-administered males. By histological observation, occurrence of spermatozoa was confirmed in scLH- and scFSH–scLH-administered eels. Although milt production was not induced, higher dosage and frequency of scGTH administration was effective in promoting testicular development of immature eels. Thus, single-chain fish GTHs produced by the baculovirus–silkworm larvae system could be a useful tool for promotion of gonadal maturation in aquaculture fishes.     

Distribution of a fish pathogen Listonella anguillarum in the Japanese flounder Paralichthys olivaceus hatchery

The present study was undertaken to investigate the distribution of Listonella anguillarum in a Japanese flounder (Paralichthys olivaceus) hatchery. A total of 2704 isolates were obtained from the developing fish, live diets and artificial feeds of Japanese flounder and their rearing water, 439 of which were identified as L. anguillarum by the combining incubation on thiosulfate-citrate-bile salt-sucrose (TCBS) agar at 35 °C overnight with polymerase chain reaction (PCR) detection for the VAH1 hemolysin gene. L. anguillarum was detected in all seven rotifer samples, with densities of 2.5 × 10³ to 4.6 × 10⁶ colony forming units (CFU) g^− 1. Both the analyzed samples of Nannochloropsis oculata contained this bacterium at densities of 1.6 × 10⁴ to 1.4 × 10⁵ CFU g^− 1. L. anguillarum was detected in only one of four samples of Artemia nauplii with a density of 4.8 × 10⁵ CFU g^− 1 (35%) and it was not detected in the two analyzed artificial feed samples. L. anguillarum was detected in 11 of 18 specimens of larval and juvenile Japanese flounder at densities of 5.0 × 10¹ to 7.4 × 10⁵ CFU g^− 1, while it was not detected in the two analyzed egg specimens of Japanese flounder. These results indicate that L. anguillarum associated with the developing Japanese flounder is likely derived from rearing water and live diets such as rotifers. Further, it is strongly suggested that L. anguillarum is a transient bacterium of the intestinal microflora for the Japanese flounder but is a permanently indigenous one for the Japanese flounder hatcheries.     

日本鳗鲡产卵后生存及其繁育

为证明日本鳗鲡(Anguilla japonica)生活史最后一步一产后鳗的命运,本研究模拟产后的日本鳗鲡继续在海水中养殖,观察其存活率及繁育情况.结果表明,产后鳗在海水中停食约18 d后,体能得到恢复,部分亲鱼开始出现摄食,1个月左右全部恢复摄食,经244 d养殖,雌、雄鳗体质量增加,存活率达94.6%.随后,给产后鳗注射外源性促性腺激素(鲤鱼脑垂体匀浆CPE和人绒毛膜促性腺激素HCG)后激发其退化的性腺(卵巢和精巢)重新发育(与当年银鳗作对照).通过性腺组织切片观察产后鳗和对照鳗性腺发育成熟的全过程及其差异,发现产后鳗起初性腺发育比当年银鳗差,但经多次注射激素后,产后鳗性腺成熟与当年银鳗同步,证明产后鳗生殖细胞对激素的敏感件高.应用17α,20β-双羟孕酮和促性腺激素释放激素类似物(GnRH-A3)使催熟的产后鳗和对照鳗均产卵和排精,并孵化出仔鱼,从而有力地证明,鳗鲡产后虽体质弱,但待体能恢复后能够继续生存和繁殖.本研究旨在探讨利用产后鳗作为今后鳗鲡人工繁殖亲鳗的可行性.     

AFLP-based genetic linkage map of marine shrimp Penaeus (Fenneropenaeus) chinensis

This paper presents the genetic linkage map of the Chinese shrimp Penaeus (Fenneropenaeus) chinensis constructed with 472 AFLP markers. A hundred F₁ progeny from an intercross between a female from the new variety “Yellow Sea No. 1” and wild caught male used for the mapping study. Two separate maps were constructed for each parent. The female linkage map consisted of 197 marker loci forming 35 linkage groups and spanned a total length of 2191.1 cM, with an average marker space of 13.5 cM. The male map consisted of 194 marker loci mapped to 36 linkage groups and spanned a total length of 1737.3 cM, with an average marker spacing of 11.0 cM. The level of segregation distortion observed in this study was 12.2%. The estimated genome length of P. chinensis was 3150.3 cM for the female and 2549.3 cM for the male, respectively. The observed genome coverage was 69.6% for the female and 68.1% for the male map. The linkage maps constructed in this study provide basic information for further linkage studies on Chinese shrimp, and more importantly, the construction of the maps are part of the work of the genetic breeding programs which will be used for growth discovered in the QTL analysis of P. chinensis.     

Assessing sexual maturity of feminized Japanese eel Anguilla japonica by measuring eye size

For efficient production of Japanese eel Anguilla japonica eggs, knowledge of the status of the sexual maturity of potential broodstock females is important because this status directly influences the time required to produce mature eggs by successive hormone doses. Here we apply an eye index (relative eye size) to evaluate the gonadal status of feminized A. japonica, which were induced by administration of estradiol-17β. Examination of gonad somatic and eye indexes of 267 feminized eels, cultivated for 12–56 months, revealed a significant correlation between these two indexes. Artificial maturation of 35 feminized eels showed that the number of hormone injections administered before ovulation was significantly lower as the eye index score increased, indicating availability of this noninvasive method of predicting sexual maturity of feminized eel. There was no correlation between eye index and fertilization rate, hatching rate, or larval survival rate, suggesting that sexual maturity before hormonal treatment does not affect egg quality.     

The roles of bone and muscle as phosphorus reservoirs during the sexual maturation of female Japanese eels, Anguilla japonica Temminck and Schlegel (Anguilliformes)

We sought to determine the organs that function as phosphorus (P) reservoirs for supplying this mineral to the ovary during the sexual maturation of the Japanese eel, Anguilla japonica (Anguilliformes). The P content of the ovary increased markedly during sexual maturation despite the lack of an extrinsic P supply (e.g., diet). This phenomenon suggests that the P that accumulated in the ovary was derived from other parts of the eel. In addition, the P content of bone dramatically decreased during maturation, as did the total bone weight. This result suggests that resorption of bone occurred during the sexual maturation of the eel. Whereas the P content and mass of muscle markedly decreased during maturation, the Ca and P contents of the skin (which was scaled) did not. The total (whole-fish) P content was constant throughout maturation. Together, our results suggest that almost all of the P lost from bone and muscle is incorporated in the ovary and that bone and muscle function as P reservoirs during the sexual maturation of the Japanese eel. This revised version was published online in August 2006 with corrections to the Cover Date.     

Video observation of an eel in the Anguilla japonica spawning area along the West Mariana Ridge

Some anguillid spawning areas are known based on collections of small larvae, but recently for the Japanese eel Anguilla japonica, adult spawners have been caught in trawls and their eggs and preleptocephali collected. The spawning area of A. japonica is located along the western side of the West Mariana Ridge, but the natural spawning behavior of this species or that of any other anguillid species has never been observed. This study reports on the first effort to observe spawning aggregations of anguillid eels that was conducted by the R/V Yokosuka using the Shinkai 6500 submersible and a Deep-Tow camera system in the A. japonica spawning area in July 2012. The submersible was deployed mostly at 200–800 m during daytime and the Deep-Tow was deployed mostly at 130–250 m during nighttime, both in multiple oblique depth tracks along linear transects. Various fishes and invertebrates were seen in the pelagic environment during day and night, but no spawning aggregations were observed. One eel was briefly recorded by a Deep-Tow camera at 20:13 on 17 July (2 days before new moon) at a depth of 179 m. The eel was recorded for <1 s as it passed in front of the camera. Its anterior body and head shape were consistent with a male A. japonica, or possibly a Derichthys serpentinus eel, but not with other mesopelagic eels. Because the tail region of the eel was not visible, species identification was not possible.     

Serotonin stimulates ovarian maturation and spawning in the black tiger shrimp Penaeus monodon

Serotonin (5-hydroxytryptamine, 5HT) has been reported to induce ovarian maturation and spawning in the crayfish Procambarus clarkii and white Pacific shrimp Litopenaeus vannamei. The aim of this study was to explore the role of exogenous 5HT on the reproductive performance of the black tiger shrimp Penaeus monodon. 5HT solution was injected into domesticated P. monodon broodstock at 50 μg/g body weight and ovarian maturation and spawning were recorded. The presence of 5HT in the ovary and oviduct of P. monodon was also studied by immunohistochemistry and its levels in the ovary by enzyme link immunoabsorbance assay (ELISA). The 5HT-injected P. monodon developed ovarian maturation and spawning rate at the level comparable to that of unilateral eyestalk-ablated shrimp. Hatching rate and the amount of nauplii produced per spawner were also significantly higher in the 5HT-injected shrimp, compared to the eyestalk-ablated shrimp. 5HT-positive reactions were found in the follicular cells of pre-vitellogenic oocytes, in the cytoplasm of early vitellogenic oocytes and on the cell membrane and cytoplasm of late vitellogenic oocytes. 5HT in the ovary was present at 3.53 ± 0.26 ng/mg protein level in previtellogenic stage and increased to 17.03 ± 0.57 ng/mg protein level in the mature stage of the ovary. The results suggest a significant role of 5HT, possibly directly on the ovary and oviduct, on the reproductive function of female P. monodon.     

Production of recombinant goldfish gonadotropins by baculovirus in silkworm larvae

Recombinant gonadotropins (GTH), follicle-stimulating hormone (FSH) and luteinizing hormone (LH) of goldfish Carassius auratus were produced by baculovirus in silkworm larvae. Hemolymph containing recombinant FSH (rFSH) or LH (rLH) was collected from silkworm larvae, and its biological activity was examined in vivo using male goldfish and female bitterling Rhodeus ocellatus ocellatus. Injection of hemolymph containing rFSH or rLH induced milt production in male goldfish, whereas only rLH induced ovulation in female bitterling. These results suggest, that biologically active goldfish recombinant GTH could be applied for the induction of gonadal development in aquaculture fishes as a substitute of pituitary extract, if a method of large scale production is established.     

Synthesis and possible function of 11-ketotestosterone during oogenesis in eel (Anguilla spp.)

The control of 11-ketotestosterone (11-KT) biosynthesis and its physiological roles were examined in female Japanese eel (Anguilla japonica) and New Zealand longfinned eel (Anguilla dieffenbachii). 11-KT was detected in serum of female eels of both species. Among various tissues from Japanese eel, the ovary had the greatest capacity to synthesize 11-KT in vitro. In addition, the oocyte diameters of eels treated with 11-KT had increased significantly. Furthermore, these oocytes were found to have an increased number of oil droplets. These findings suggest that 11-KT in female eels may be mostly of ovarian origin and that this androgen appears to play an important role in controlling pre-vitellogenic oocyte growth.     

Development of an in vitro culture system for producing eel larvae from immature ovarian follicles in Japanese eel <Emphasis Type="Italic">Anguilla japonica</Emphasis>

To standardize conditions during the final maturation and ovulation of ovarian follicles from Japanese eel, we have developed a culture system for the production of fertilizable eggs from post-vitellogenic ovarian follicles in vitro. Post-vitellogenic ovarian follicles were incubated in culture medium supplemented with 17α,20β-dihydroxy-4-pregnen-3-one (DHP) with or without bovine serum albumin (BSA) to assess the effects of protein concentration. Eggs that ovulated during incubation were fertilized, and the remaining follicles were incubated in prostaglandin F_2α (PGF_2α) for a further 3 or 6 h before fertilization. Male eels were injected repeatedly with human chorionic gonadotropin. The quality of eggs obtained under the different culture conditions was evaluated after artificial fertilization in terms of hatching success. Hatching rates tended to decrease with increasing concentrations of BSA in the incubation medium in a dose-dependent manner. The addition of PGF_2α drastically increased the number of eggs that ovulated, but the rate of hatching was greatly decreased compared with eggs obtained earlier by DHP incubation alone. The larvae obtained from artificially fertilized eggs produced in vitro survived for 14 days without feeding. We conclude that in vitro culture systems thus have a great potential for the acquisition of good quality eggs under tightly controlled artificial conditions, culminating in the production of eel larvae.     

Changes in the immunostaining intensities of follicle-stimulating hormone and luteinizing hormone during ovarian maturation in the female Japanese flounder

The role of gonadotropin (GTH) in the reproduction of the Japanese flounder, Paralichthys olivaceus, was studied by assessing the changes in the apparent activity of follicle-stimulating hormone (FSH) and luteinizing hormone (LH) in the pituitary gland during gonadal maturation by immunohistochemical analyses. Corresponding changes in plasma levels of testosterone (T), estradiol-17β (E₂), and 17α,20β-dihydroxy-4–pregnen-3-one (DHP) were also studied. Reared fish at the early spawning to termination stages were sampled from May to August and wild fish at the previtellogenic to termination stages were caught at 3- to 4-week intervals between April and September offshore from the northern mainland of Japan by gill nets. The gonadosomatic index of the reared fish decreased from the early spawning stage to the termination stage, while that of the wild fish increased significantly from the previtellogenic stage to the early spawning stage and decreased thereafter. In the reared fish, the immunostaining intensities of FSH and LH were high during the spawning period, accompanied by high plasma levels of T, E₂, and DHP. In the wild fish, the immunostaining intensities of FSH and LH were low during the previtellogenic stage but increased during the maturing and spawning stages. These results indicate that both FSH and LH are likely associated with oocyte maturation in the Japanese flounder.     

Effects of two dietary probiotics (Bacillus subtilis or licheniformis) with two prebiotics (mannan or fructo oligosaccharide) in Japanese eel,Anguilla japonica

A 12‐week feeding trial was conducted to investigate the effects of two dietary probiotics; Bacillus subtilis KCTC 2217 or Bacillus licheniformis KCCM 11775 with two prebiotics; mannan oligosaccharide (MOS) or fructooligosaccharide (FOS) in Japanese eel. Fish averaging 12.8 ± 0.47 g (mean ± SD) were randomly distributed into five treatments with triplicate tanks. A basal control diet (CON) and four synbiotic diets supplementing B. subtilis + MOS (BSM), B. subtilis + FOS (BSF), B. licheniformis + MOS (BLM), and B. licheniformis + FOS (BLF). Weight gain and specific growth rate of fish fed all synbiotic diets were higher than those of fish fed CON. Immune‐related gene expression of heat shock protein 70 and immunoglobulin M of fish fed BSF and BLM were significantly higher than those of fish fed CON. Fish fed BSF and BLM had significantly higher intestinal villi length than those of fish fed BLF and CON (p < .05). Disease resistance against Aeromonas hydrophila of fish fed all synbiotic diets were significantly higher than those of fish fed CON (p < .05). Therefore, these results indicated that dietary B. subtilis with FOS (BSF) and B. licheniformis with MOS (BLM) could have beneficial effects on intestinal morphology, and immune‐related gene expression in Japanese eel.     

Sexual dimorphism of fast muscle fibre recruitment in farmed Atlantic halibut (Hippoglossus hippoglossus L.)

Commercially farmed Atlantic halibut (Hippoglossus hippoglossus) were reared at the Aga Marin's facility located on Dønna (Norway) under ambient environmental conditions in duplicate 15 × 15 × 8 m netpens (May 2004 to May 2005). Twenty fish were sampled five times over a twelve month production period during which time the average body weight increased from 1.26 to 2.08 kg (n = 100 fish). Body mass, fork length (L_F), and the number and size distributions of fast muscle fibres were determined in male and female fish. All males matured during the autumn whereas no maturation was observed in females. From the point of maturation females had superior growth performance to males and body mass and the total cross-sectional area of muscle were 1.4-fold (P < 0.05) and 1.3-fold higher (P < 0.01) respectively by May 2005. The total number of fast muscle fibres per trunk cross-section at 0.55 L_F was 24.5% higher in females (7.58 × 10⁵) than males (5.80 × 10⁵) prior to sexual maturation. In females, muscle fibre recruitment slowed with short days and low water temperatures (< 6.5 °C), but had increased to 1.01 × 10⁶ by May 2005 (P < 0.001). In contrast, there was no growth and no increase in muscle fibre number in males following the onset of maturation. The distribution of muscle fibre diameters prior to maturation in males was also significantly different between male and female fish matched for L_F. The results illustrate a sexual dimorphism of muscle fibre recruitment patterns in Atlantic halibut and highlight the adverse affects of sexual maturation in males on muscle growth.     

11-ketotestosterone potentiates estrogen-induced vitellogenin production in liver of Japanese eel (Anguilla japonica)

Isolated hepatocytes from male Japanese eel at various stages of human chorionic gonadotropin (HCG)-induced spermatogenesis were cultured in the presence of estradiol-17β (E2). Hepatic vitellogenin (VTG) production in response to E2 increased during testicular development. This indicates that VTG production is influenced by gonadal maturity not only in females but also in males, where serum levels of 11-ketotestosterone (11KT) increase during testicular development. Recently, it has been confirmed that significant levels of 11KT are detected in maturing female eel. Therefore, in a next experiment, effect of 11KT on E2-induced VTG production was examined in vitro. As a result, in both male and female hepatocytes, 11KT enhanced E2-induced VTG production, although 11KT alone failed to induce VTG production.     

Immunohistochemical changes in production of pituitary hormones during artificial maturation of female Japanese eel <Emphasis Type="Italic">Anguilla japonica</Emphasis>

ABSTRACT:   Specific antibodies against follicle-stimulating hormone β subunit (FSHβ), prolactin (PRL), and somatolactin (SL) of the Japanese eel Anguilla japonica were produced. These antibodies, as well as antibodies against luteinizing hormone β subunit (LHβ) and growth hormone (GH) produced previously, were used to examine changes in the production of pituitary hormones in female eels during maturation induced by salmon pituitary homogenate (SPH) injection. Immunohistochemical observations showed a decrease in FSH production after SPH injection, suggesting that SPH inhibits FSH production. In contrast, LH production increased markedly with maturation. The number of GH producing cells decreased gradually during maturation, possibly because of inhibition by exogenous GH present in the SPH and/or endogenous insulin-like growth factor-I produced by the stimulation of salmon GH. Although changes in the number of PRL producing cells with maturation were not evident, the number of SL producing cells showed a peak at the late vitellogenic stages, and thereafter decreased to the migratory nucleus stage. These results suggest that GH and SL are involved in sexual maturation in SPH injected eels, as in other fishes.     

Estimation of production and long‐term changes in the population of European eel Anguilla anguilla (L.) in an Irish lake

Monitoring of yellow‐phase and silver‐phase Anguilla anguilla during their continental life history is necessary for evaluation of stock recovery measures. Eel population data for an Irish lake (Lough Sheelin) were compiled for the period 1993–2014. Catch data from 2009 to 2014 provided minimum estimates of recent silver eel production ranging annually from 0.79 to 1.84 kg/ha. Long‐term changes in yellow eel abundance and silver eel size structure were assessed as part of a fishery monitoring programme. Yellow eel catch per unit effort (CPUE) in the lake (from longline surveys) was considerably higher in the 1990s (52.2–62.1 eels/100 hooks) than 2002–2008 (1.9–15.8 eels/100 hooks). Conversely, during 1993–2014, the mean size of silver eels migrating from the lake increased significantly (p < .001), from 659 mm to 838 mm. The results suggest that in the absence of direct yellow eel abundance data, interannual variation in silver eel size structure may be a useful monitoring tool for local eel stocks as part of Eel Management Plans (EMP's).