During the long migration from river habitats to the spawning ground, the Japanese eel undergoes sexual maturation. This spawning migration occurs concurrently with morphological changes, such as increases in eye size; however, the mechanisms by which sex steroids and their receptors influence these changes in peripheral tissues remain unclear. The aim of this study was to investigate changes in the eyes of female Japanese eels during sexual maturation, and our research focused on estrogen receptor (ER)α and ERβ transcripts. During ovarian development, the gonadosomatic index increased and yolk-laden oocytes developed rapidly. These changes occurred in conjunction with a steady increase in plasma levels of estradiol-17β (E₂). Concomitant increases in transcript levels of ERα and ERβ in eye, brain, pituitary, and ovary were also observed. Fluorescence in-situ hybridization analyses revealed that ERα and ERβ transcripts were present in the choriocapillary layer and photoreceptor layer of the eyes, and the analysis also revealed that their signals in these layers became stronger in mature females compared to those observed in immature females, suggesting that under the influence of gonadotropins, morphological changes in the eyes are regulated by E₂ through the activation of its receptors. In conclusion, E₂ plays a crucial role in physiological adaptations that occur in peripheral tissues during the spawning migration.

The freshwater eel Anguilla japonica is rapidly decreasing in number and has not yet been successfully mass produced. This may be at least partially attributable to the unique and long early life history of the eel. Therefore, we investigated its ontogeny of morphometry and growth pattern in larval stages to provide baseline information for understanding the early life history and improving seed rearing technology. This study was conducted for 200 days after hatching (DAH) and analyzed morphometry and allometry for eel larvae. The following cultured eel larval stages were identified: the yolk sac larvae stage (0–6 DAH, 3.23–6.85 mm total length (TL)), the pre-leptocephalus stage (7–30 DAH, 6.85–15.31 mm TL), and the leptocephalus stage (50–200 DAH, 15.31–60.06 mm TL). Cultured and wild eel larvae could be divided into characteristic larval stages at similar sizes. However, compared to wild eels, cultured eels had a slower growth rate and fewer preanal myomeres. Meanwhile, cultured eel larvae rarely had a mixed feeding period as the absorption of endogenous reserves was completed by 7 DAH. The lower jaw of eel larvae was significantly longer than the upper jaw from 50 DAH. In the pre-leptocephalus and leptocephalus stages, eel larvae showed continuous positive allometric growth at trunk height and tail muscle height with change to the willow leaf-like form. These growth characteristics may be the result of adaptation to the migration over long distances and to a diel vertical migration. The inflection point in the body parts growth patterns showed only before 30 DAH, and mass mortality appeared at this period. Therefore, to improve the growth and survival rates of cultured eel seed, it is necessary to focus on improving the feeding and rearing protocol until 30 DAH.

This study aimed to produce single-chain recombinant Anguillid eel follicle-stimulating hormone (rec-eel FSH) analogs with high activity in Cricetulus griseus ovary DG44 (CHO DG44) cells. We recently reported that an O-linked glycosylated carboxyl-terminal peptide (CTP) of the equine chorionic gonadotropin (eCG) β-subunit contributes to high activity and time-dependent secretion in mammalian cells. We constructed a mutant (FSH-M), in which a linker including the eCG β-subunit CTP region (amino acids 115–149) was inserted between the β-subunit and α-subunit of wild-type single-chain eel FSH (FSH-wt). Plasmids containing eel FSH-wt and eel FSH-M were transfected into CHO DG44 cells, and single cells expressing each protein were isolated from 10 and 7 clones. Secretion increased gradually during the cultivation period and peaked at 4000–5000 ng/mL on day 9. The molecular weight of eel FSH-wt was 34–40 kDa, whereas that of eel FSH-M increased substantially, with two bands at 39–46 kDa. Treatment with PNGase F to remove the N glycosylation sites decreased the molecular weight remarkably to approximately 8 kDa. The EC₅₀ value and maximal responsiveness of eel FSH-M were approximately 1.23- and 1.06-fold higher than those of eel FSH-wt, indicating that the mutant showed slightly higher biological activity. Phosphorylated extracellular-regulated kinase (pERK1/2) activation exhibited a sharp peak at 5 min, followed by a rapid decline. These findings indicate that the new rec-eel FSH molecule with the eCG β-subunit CTP linker shows potent activity and could be produced in massive quantities using the stable CHO DG44 cell system.

We investigated the nine experimental diets containing fish meal (FM) and/or fish meal analog (FMA) as the major source of animal protein to determine the optimum FMA level as the substitute of FM protein in the diet of juvenile Japanese eel. In addition, two natural feed additives such as Song-Gang stone (SG) and Yucca meal (YM) were supplemented in the diet to evaluate their efficacy as the immunostimulants. The diets are as follows: 100% FM + 0% FMA in diet (FMA₀), 90% FM + 10% FMA in diet (FMA₁₀), 80% FM + 20% FMA in diet (FMA₂₀), 70% FM + 30% FMA in diet (FMA₃₀), 60% FM + 40% FMA in diet (FMA₄₀), FMA₀ + 0.4% SG (FMA₀SG), FMA₀ + 0.1% YM (FMA₀YM), FMA₂₀ + 0.4% SG (FMA₂₀SG), and FMA₂₀ + 0.1% YM (FMA₂₀YM). Nine groups of Japanese eel each with three replicates were distributed (initial weight of 9 ± 0.2 g) in rectangular tanks receiving flow through water. Each group of the treatment consisted with 15 fish and fed one of the diets for 8 weeks. At the end of the feeding trial, fish fed with the FMA₀ and FMA₁₀ diets showed no significant differences in weight gain (WG), specific growth rate (SGR), feed efficiency (FE), and protein efficiency ratio (PER). Meanwhile, fish fed with FMA₂₀, FMA₃₀, and FMA₄₀ diets showed significantly lower WG, SGR, FE, and PER than the fish fed with the FMA₀ (control) diet. In addition, there were no significant differences among fish fed with the SG- and YM-supplemented diet groups. However, lysozyme activities in fish fed with the FMA₁₀, FMA₂₀, FMA₃₀, and FMA₄₀ were significantly lower than the fish fed with the FMA₀SG, FMA₀YM, FMA₂₀SG, and FMA₂₀YM diets. After 7 days of injection with V. Anguillarum, cumulative survival rates of fish fed with the FMA₀SG and FMA₀YM diets were significantly higher than the FMA₀ diet group. The results revealed that the FMA could replace up to 10% of FM as a protein source in the diet of Japanese eel and both of the natural feed additives (SG and YM) could improve replacing rates of FMA from 10% to 20% without compromising growth and health status of fish.

Beneficial microorganisms can increase nutrient digestion and absorption in farmed fish. This study investigates the effects of supplemental feeding of Bacillus species isolated from the intestines of wild glass eels on the growth, survival, and gene expression of farm-raised eel larvae for 30 days after hatching. Three species of Bacillus (B. velezensis, AJBV; B. subtilis, AJBS; B. licheniformis, AJBL) without hemolytic activity were isolated, and an experiment compared the growth of eel larvae fed an artificial diet supplemented with each Bacillus species. There were no significant differences in the total length and body depth of eel larvae at 30 days after hatching in all groups. During the feeding period, 149 eels survived from the initial 1000 in the control group. On the other hand, 240, 178, and 141 eels survived in the AJBV, AJBS, and AJBL groups, respectively. However, there were no significant differences in survival rates despite the difference in the number of surviving eel larvae among the groups. In the comparison of gene expression of genes involved with growth (growth hormone, growth hormone receptor 1, insulin-like growth factor II-2) and those involved with digestive enzymes (amylase, trypsin, lipase), there were also no significant differences among the groups. Our results confirm that dietary supplementation with each of the three host-associated Bacillus does not affect the growth and survival rates of eel larvae reared on an artificial diet up to the first 30 days after hatching, nor does it significantly affect related gene expression.