Japanese Eel Anguilla Japonica Research Articles

Asymmetric competition for habitats between the temperate Japanese eel Anguilla japonica and the tropical Indo-Pacific eel A. marmorata

Habitat segregation is a common phenomenon among anguillids in rivers where multiple species coexist. However, despite the growing need to conserve their river habitats, the mechanisms underlying such segregation remain unknown. Here, we conducted tank experiments to investigate the competitive superiority for habitats between the temperate Japanese eel Anguilla japonica and the tropical Indo-Pacific eel Anguilla marmorata, which co-occur in East Asian subtropical rivers. Using eight interspecific pairs of these two species of comparable total length, we compared the use of a single pipe shelter, recorded over a 24-h experimental period, between solitary and paired conditions. The changes in shelter use patterns between conditions indicated that in the paired condition, the shelter was occupied by A. marmorata in five pairs (62.5%) by the end of the experiment while A. japonica appeared to occupy the shelter in only one pair (12.5%). Furthermore, agonistic behaviors, which occurred mainly at night, such as biting and head-butting, were observed much more frequently for A. marmorata toward A. japonica (mean ± standard deviation: 23.2 ± 36.6 times per individual) than for A. japonica toward A. marmorata (0.4 ± 0.5 times per individual), indicating an asymmetry in aggressiveness. These results suggest that A. marmorata can negatively affect the diurnal habitat use and nocturnal activities of sympatric A. japonica. These findings, combined with those from previous field studies, suggest that A. marmorata may competitively exclude A. japonica from stream habitats with large substrate sizes. This asymmetric competition can contribute to shaping their riverine distribution and composition within rivers where they co-occur.

How Japanese eels escape from the stomach of a predatory fish

Predation shapes diversity in the defensive tactics of prey. One specialized defensive tactic is to escape the digestive system of the predator after capture1,2,3,4,5,6,7,8. While most of these defensive tactics involve passive ejection alive from predators' mouths and vents1,2,3,4,5, active escape from the digestive tracts of predators has recently been observed in certain invertebrate species6,7 and fish8. However, no study has yet uncovered the behavioral patterns and escape routes of the prey within a predator's digestive tract. Here, we report the sequential escape processes of the Japanese eel Anguilla japonica from capture to escape via the gills of predatory fish Odontobutis obscura using an X-ray video system. All captured eels had at least one portion of their bodies swallowed into the stomach of the predator. Surprisingly, after being swallowed, most individuals attempted to escape by going back up the digestive tract towards the esophagus and gill, and some of them succeeded in escaping via the predator's gill. Some eels, whose entire bodies were completely inside the stomach, exhibited circling behavior along the stomach, seemingly searching for possible escape routes. An electro-anesthetization experiment revealed that eels utilize various escape routes through gill clefts, rather than just one.

Public preferences for policies promoting the conservation of a universally threatened species (Anguilla spp.): Insights from a choice experiment in Japan

The population of the Northern temperate eel species (Anguilla spp.) has undergone a significant decline, resulting in their classification as Endangered (American eel Anguilla rostrata and Japanese eel Anguilla japonica) or Critically Endangered (European eel Anguilla anguilla) by the IUCN. The growing global demand for Japanese cuisine and processed eel products could potentially exacerbate the vulnerability of these species. This research examines the acceptability of conservation policies among Japanese consumers. A discrete choice experiment was employed to investigate the social acceptance of target levels for national eel consumption reduction and to examine the trade-offs made by individuals between various policy instruments (e.g., combat against illegal trade, research on artificial reproduction, shift to substitute species, public awareness campaigns, eco-labelling) and the costs that they may be willing to incur. The econometric analysis identified two distinct groups with contrasting preferences. The first group exhibited strong support for the current situation. Conversely, the second group expressed preferences for expanding public intervention. We further explored the sources of heterogeneity. Paradoxically, we found that education attainment and environmental knowledge in relation to consumption motivations influenced individual preferences for the status quo. However, individuals with higher pro-environmental attitudes are more likely to accept a national target of consumption reduction. They are willing to pay for demand-oriented policy instruments to complement existing supply-side measures.

Microbial influence on the larval survival of Japanese eel Anguilla japonica: Antibiotic-mediated alterations and biomarker isolation.

In rearing systems for the Japanese eel Anguilla japonica, although it is assumed that microorganisms influence larval survival and mortality, particularly during the early stages of growth, the effects of bacterial communities on larval survival have yet to be sufficiently determined. In this study, we compared the bacterial communities associated with larval survival at three stages of eel growth. To artificially alter bacterial communities and assess larval survival, eel larvae were treated with 11 types of antibiotic, and larval survival and bacterial characteristics were compared between the antibiotic-treated and antibiotic-free control groups. Throughout the three growth stages, eels treated with four antibiotics (polymyxin B, tetracycline, novobiocin, and erythromycin) had survival rates higher than those in the control groups. The bacterial communities of surviving larvae in the control and antibiotic groups and dead larvae in the control groups were subsequently analyzed using 16S rRNA gene amplicon sequencing. PERMANOVA analysis indicated that these three larval groups were characterized by significantly different bacterial communities. We identified 14 biomarker amplicon sequence variants (ASVs) of bacterial genera such as Oceanobacter, Alcanivorax, Marinobacter, Roseibium, and Sneathiella that were enriched in surviving larvae in the antibiotic treatment groups. In contrast, all four biomarker ASVs enriched in dead larvae of the control groups were from bacteria in the genus Vibrio. Moreover, 52 bacterial strains corresponding to nine biomarkers were isolated using a culture method. To the best of our knowledge, this is the first study to evaluate the bacterial communities associated with the survival and mortality of larvae in during the early stages of Japanese eel growth and to isolate biomarker bacterial strains. These findings will provide valuable insights for enhancing larval survival in the eel larval rearing systems from a microbiological perspective.

The effects of two dietary synbiotics on growth performance, hematological parameters, and nonspecific immune responses in Japanese Eel.

Feed additives have attracted increased attention in aquaculture due to their ability to modulate fish gut microbiota, resulting in improved fish growth and immunity. This study assessed the effects of two synbiotics in Japanese Eel Anguilla japonica: Bacillus subtilis with mannooligosaccharides (MOS) and Enterococcus faecium with fructooligosaccharides (FOS). Six diets, including a control (CON) diet, oxytetracycline (OTC) diet, and four synbiotic diets (B. subtilis at 1 × 106 or 1 × 107 colony-forming units [CFU]/g with MOS at 5 g/kg [BS6MO and BS7MO; collectively, BSMOS diets] and E. faecium at 1 × 106 or 1 × 107 CFU/g with FOS at 5 g/kg [EF6FO and EF7FO; collectively, EFFOS diets]), were fed to triplicate groups of 20 fish (average weight ± SD = 6.00 ± 0.07 g) for 8 weeks. Fish fed the BSMOS diets showed significantly higher weight gain, specific growth rate (SGR), and feed efficiency compared to fish fed the CON and OTC diets, but the values were not significantly different from those of fish fed the EFFOS diets. Weight gain and SGR of fish that were given EFFOS diets were not significantly different from those of fish fed all other diets. Fish fed the OTC diet showed a higher mean aspartate aminotransferase level, although the difference was not statistically significant. The myeloperoxidase activity of fish fed the BS7MO diet was significantly higher than those of fish receiving all other diets, and the superoxide dismutase activity of fish fed the BS7MO diet was also significantly higher than that of fish fed the EF7FO diet. Overall, the BSMOS synbiotic diets were significantly more effective than the CON diet in enhancing fish survival against a Vibrio anguillarum challenge. Our findings suggest that synbiotics can be a preferable alternative to antibiotics in aquaculture.

Diets comprising hen egg yolk and milk proteins as potential alternatives to shark egg-based diets for larvae of the Japanese eel Anguilla japonica

Diets comprising hen egg yolk and milk proteins as potential alternatives to shark egg-based diets for larvae of the Japanese eel Anguilla japonica

Effects of Hot-Spring Water Culture on the Nutritional Value of Muscle from Japanese Eel Anguilla japonica

ABSTRACT The chemical composition of muscles from Japanese eels cultured in hot-spring water was determined and compared with that of eels cultured in well water. The proximate composition did not differ significantly between the culture conditions, but the hepatosomatic indices of the hot-spring water eels were significantly higher than those of the well water eels. Amino acid levels were higher in the hot-spring water eels, and the carnosine content of the hot-spring eels was lower than that of the controls. Our results suggest that hot-spring water is a promising medium for eel culture, and its use may bring energy cost savings.

Effects of luteinizing hormone-releasing hormone analog and pimozide on the release of luteinizing hormone and ovulation in artificially matured Japanese eel Anguilla japonica

Pituitary gonadotropins, follicle-stimulating hormone (Fsh) and luteinizing hormone (Lh), control oogenesis in all vertebrates. In particular, Lh plays a key role in stimulating the final oocyte maturation and subsequent ovulation. The biosynthesis and secretion of Lh are regulated by several neurohormones, including gonadotropin-releasing hormone (GnRH) and dopamine. GnRH analogs, also known as Lh releasing hormone analogs (LHRHa), and dopamine antagonists are commonly used to induce sexual maturation in teleosts. However, the effects of these reagents differ among fish species. Therefore, in the present study, the effects of LHRHa and pimozide (a dopamine antagonist) on Lh release and ovulation induction were investigated in female eels whose ovarian development was artificially induced by recombinant Fsh with ovaries containing oocytes at the migratory nucleus stage in vitro and/or in vivo. Both LHRHa and pimozide stimulated the release of Lh from pituitary cells in a dose-dependent manner in vitro. Furthermore, the synergistic effects of LHRHa and pimozide were observed in the release of Lh from the pituitary gland. In vivo experiments demonstrated that the administration of pimozide alone or in combination with LHRHa induced the release of Lh and the fusion of oil droplets in oocytes. In addition, 17α-hydroxyprogesterone (OHP) combined with LHRHa and pimozide resulted in complete ovulation at a high rate (87.5% complete ovulation and 12.5% partial ovulation), whereas LHRHa and pimozide without OHP induced partial ovulation at a low rate (0% complete ovulation and 14.3% partial ovulation). These results suggest that combined treatment with LHRHa, pimozide, and OHP could induce ovulation in artificially matured female Japanese eels. However, future studies are needed to evaluate the quality of embryos produced by females induced with LHRHa, pimozide and OHP. Additionally, the pituitary gland obtained from female eels with artificially induced sexual maturation is suitable for investigating the mechanisms of Lh release.

Ontogenetic habitat shift of Anguilla marmorata and A. japonica in the rivers of southern Japan: Implications for habitat use evaluation and conservation

Abstract Understanding the habitat requirements of fishes throughout their life cycle is essential for effective conservation. Despite the growing need to conserve anguillid eels, the current understanding of the ontogenetic habitat shifts during their riverine stages remains limited, particularly for the Japanese eel Anguilla japonica and the Indo‐Pacific eel Anguilla marmorata. Here, changes in habitat use during the riverine stages of these two eel species were investigated in the southern Japan rivers, where they coexist. Eels were classified into total length (TL) classes at 10 cm intervals, and generalized additive models were constructed using these classes as an ordinal, quasi‐continuous explanatory variable. The models showed that the habitat use of the two species changed with growth; both species shifted to deeper and more lentic habitats as they grew larger. Small‐ and mid‐sized (<25 and 25–55 cm in TL, respectively) A. japonica used more lentic habitats with smaller substrates, ranging from sand/silt to cobbles, than small‐ and mid‐sized A. marmorata, which mainly used lotic habitats with cobble and boulder substrates. However, compared with small‐sized conspecifics, mid‐sized A. japonica mainly used habitats with larger substrates, such as pebbles and cobbles, suggesting that the interspecific overlap in habitat use is relatively high for mid‐sized individuals of both species. Large A. marmorata ( 85 cm) mainly used habitats with smaller substrates as they grew larger. The results suggest that using a fine body size classification and including these classes as a quasi‐continuous explanatory variable in habitat models capture gradual changes in habitat use, improving habitat use evaluation of anguillid eels. Preserving the environmental heterogeneity of river habitats in terms of water depth, current velocity and substrate size would facilitate habitat segregation of both conspecifics and allospecifics in the same rivers, thereby promoting the conservation of anguillid eels.

Individual variation in post-metamorphic changes in feeding incidence, digestive organ tissues and enzyme gene expression in Japanese eel Anguilla japonica glass eels

Temporal changes of feeding incidences, digestive organ tissues, and mRNA expression of digestive enzymes were investigated in artificially reared Anguilla japonica glass eels. Incidences of daily feeding and cumulative first feeding exceeded 50% at day 40 and 33, respectively, but tended to increase gradually. Time until first feeding varied from day 11–61 among individuals. Glass eels without food after metamorphosis showed notable developmental changes in their digestive system that included increases in esophagus goblet cells and blood vessel diameters, elongation of the stomach, differentiation and increased number of gastric glands, and increases in gall bladder sizes. A distinct qualitative change, which was gastric gland differentiation, occurred in all eels until week 2 after metamorphosis. All quantitative character values showed trends of gradual changes in their averages and had high coefficients of variation, especially for goblet cell numbers. Few deaths and no histological features related to severe starvation such as notably more hepatocyte necrosis or desquamation of intestinal epithelial cells were observed even after 9 weeks. Hepatocyte-vacuole stored glycogens and pancreatic zymogen granules were also found, suggesting that they were not expressing symptoms of starvation. Relative expressions of five digestive enzymes were lowest at week 0 and increased gradually in their averages with the passage of weeks, along with large variations among fish. This study found that half of the post-metamorphic glass eels began feeding after 4 weeks, but that there were large individual variations in the timing of feeding onset, the degree of developmental of their digestive organs, and in digestive enzyme expressions.

Regional differences in oceanic migratory behavior of Japanese silver eel in waters with different vertical temperature gradients

BackgroundThe Japanese eel Anguilla japonica has a single spawning area on the west coast of the Mariana Islands, and the larvae disperse widely via the North Equatorial Current and the Kuroshio Current to freshwater and coastal areas of East Asia for nursery and growth habitats. Spawning adults reared in their respective habitats begin their migration to spawn from September to February, but little is known about regional differences in migration routes, their contribution to spawning populations, or their migratory behavior. To better understand the spawning migrations of Japanese eels, we tracked adult migratory stage eels (silver eels) captured in coastal areas of three regions (Tokai region, Sea of Japan coastline, and Tohoku region) from October 2019 to February 2020 using pop-up satellite archival tags.ResultsTwenty-three tagged eels were released in each coastal area, and data were collected on 15 eels for a maximum of 59 days. Eels released in the Tokai region migrated southeast along the Kuroshio Current. On the other hand, eels on the Sea of Japan coastline and in the Tohoku region tended to migrate westward and southward against the ocean current, respectively, but no eels from either region reached waters near the Kuroshio Current. In addition, the present study found region-specific vertical movement patterns among the eels. Eels from the Tokai region showed typical diurnal vertical movements, while an eel from the Sea of Japan coastline repeated non-diurnal vertical movements, which was attributed to regional differences in bathymetry and water mass structure. Except for one eel suspected of being preyed upon, all other eels did not venture into depth zones with temperatures below 4 ^{circ }textrm{C}, thus suggesting that the maximum depth of vertical movement is limited by water temperature.ConclusionsThis study provided new insights into the migratory behavior of Japanese silver eel from the Sea of Japan coastline and the Tohoku region, but further long-term tracking is required to reveal the fate of the spawning adults from the coast of Japan.

Novel tissue thickening around the notochord sheath found in deformed Japanese eel Anguilla japonica leptocephali.

Even though reared leptocephalus larvae of the Japanese eel Anguilla japonica have a high incidence of notochord deformities (>60%), the cause is unknown. We performed histological examinations of the notochord and associated organs in reared larvae to better understand the process causing notochord deformation in eel larvae. In deformed larvae, unknown tissue thickening was discovered near the notochord sheath. Azan staining revealed that these tissue thickenings are most likely collagen fibers within fibrous connective tissue. This was almost identical to the connective tissue found in the primordium of the vertebral body around the notochord sheath in properly metamorphosing larvae. Furthermore, the amount of the thyroid hormone triiodothyronine (T3) was significantly higher in deformed larvae than in normal larvae, indicating that notochord deformity is probably linked to metamorphosis despite the immature stage of growth. We suggest that the aberrant growth of connective tissue surrounding the notochord sheath induced by incomplete metamorphosis causes deformities in eel larvae. The reason why deformed larvae have greater thyroid hormone levels is still unknown. It is important to assess how environmental and dietary factors affect the thyroid hormone levels of eel larvae raised in captivity.

Habitat selection behaviour of Pungitius kaibarae (Gasterosteidae) in the presence of the visual predator Anguilla japonica (Anguillidae)

Our study deals with how the short nine-spine stickleback Pungitius kaibarae uses visual cues to select among habitats, thereby reducing predation pressure from the Japanese eel Anguilla japonica. We examined the behaviour of P. kaibaraein selecting four habitats (water weed, sandy, rocky and open water) under the presence/absence of A. japonica. Using video tracking technology, we found that in the presence of A. japonica, sticklebacks preferred specific refuge patches, such as waterweed and rocks, over open water with a sandy bottom. Thus, our investigation demonstrated the importance of habitat structures in evoking escape behaviour. We discuss the importance of this bold behaviour patterns of the prey fish.Keywords: Bold behaviour, Stickleback, Eel, Swimming speed, Video tracking

Active swimming and transport by currents observed in Japanese eels (Anguilla japonica) acoustically tracked in the western North Pacific

The mechanisms of oceanic animal migration remain enigmatic. Adult Japanese eels start their long-distance oceanic migration from coastal areas to breed near the West Mariana Ridge. We tracked acoustically tagged eels released in the Kuroshio Current (KC) area near Japan (five silver-phase eels, three of which had impaired swim bladders) and a tropical/subtropical (TS) area near/in the spawning area (two yellow-phase and three silver-phase eels). We analyzed their active swimming and transport by water currents. The strong flow of the KC dominated the eels’ movements in the north, and TS area; their swimming influenced their movements. In the KC area, greater distances were covered at night than during the day, because eels swam in shallower layers with strong currents at night. Three and one eel in the TS and KC area in the upper 400 m showed counterclockwise and clockwise movements around the time of solar culmination, respectively. The meta-analysis showed that eels released at middle latitudes (20°–34° N) generally swam southward through currents, whereas those released at low latitudes (12°–13° N) generally swam northward through currents. Our study suggests the influence of the surrounding current and a potential effect of solar cues on the movements of Japanese eels.

North Equatorial Current and Kuroshio velocity variations affect body length and distribution of the Japanese eel Anguilla japonica in Taiwan and Japan

The larval stage of Japanese eel travels a substantial distance over a long duration through the North Equatorial Current (NEC) and the Kuroshio, and the spawning behavior of mature eels leads to monthly arrival waves in eastern Taiwan between November and February. The total length (TL) of the glass eel relates to its larval duration and age; therefore, the TL can indicate the larval duration. The monthly mean TLs of eels along eastern Taiwan from 2010 to 2021 were used to estimate the batch age, and the recruitment patterns and relative abundances were compared. The TLs of glass eels followed a normal distribution, and the estimated ages were highly correlated with their mean TLs. Early recruit TLs were significantly greater than those of late recruits. The mean tracer drift time was longer in early recruitment months (November–December) than in later dates (February–March). The recruitment lag between Taiwan and Japan was approximately 1–1.5 months, with relative more abundance in Taiwan for the early recruits and in Japan for the late recruits. Speculated cohorts followed the main streams of the NEC and Kuroshio, and the monthly velocity changes of these currents could affect the mean TLs as well as the distribution patterns of Japanese glass eels in Taiwan and Japan.

Observation of the feeding behaviour of reared Japanese eel Anguilla japonica leptocephali fed picocyanobacteria Synechococcus spp.

The authors observed the feeding behaviour of artificially reared Japanese eel Anguilla japonica leptocephali, 7.5-19 mm total length (10-61 days post-hatch), fed Synechococcus sp., which is considered a potential food source of anguilliform larvae. Three strains of Synechococcus sp. (NIES-972, 976 and 979) were tested as the food material. Larvae across the entire length range could effectively ingest a suspension of pico-sized cyanobacteria (1-3μm in diameter). Video observations of the mid-hindgut of larvae under an epifluorescence microscope confirmed that the movement of microvilli of the intestinal epithelium allowed the cell particles to circulate in the mid-hindgut, before becoming solidified in the anal region. Significant differences in food intake were observed between larvae fed two strains of Synechococcus (NIES-972 and 976), and among different cell densities, which suggests feeding selectivity and density dependence. Comparisons of feeding behaviour under the light group (9L:15D) and the dark group (24D) showed significantly higher food intake (measured as an index of intestinal fullness) in the light group, although substantial and continuous ingestion was observed in the dark group, indicating continuous feeding by swallowing sea water. The authors hypothesise that the feeding ecology of anguilliform leptocephali is based on a survival strategy whereby the larvae do not compete with various higher-trophic-level fishes for food in an oligotrophic environment.

Characterization of ovarian culture in vitro and sex steroids in vivo by recombinant eel gonadotropin treatments in the eel Anguilla japonica

In the present study, we investigated the effects of recombinant eel gonadotropins (rec-GTHs) on maturation induction in immature ovarian culture in vitro and sex steroid hormones in vivo in the Japanese eel Anguilla japonica. To study the in vitro effects of rec-GTHs on estradiol-17β (E2) production in immature ovarian tissues, ovarian tissues were incubated with different doses of rec-follicle-stimulating hormone (rec-FSH) or rec-luteinizing hormone (rec-LH). The results revealed that the E2 levels in the rec-FSH (0.1, 0.5, or 1 μg/mL)- and rec-LH (0.1 or 0.5 μg/mL)-treated groups were significantly higher than those in the female eels from the control group. Furthermore, to investigate the in vivo effects of rec-GTHs on the gonadosomatic index (GSI) and plasma sex steroid hormone levels, the eels were injected intraperitoneally with eel’s ringer (control), salmon pituitary extract (SPE; for female eels), human chorionic gonadotropin (hCG; for male eels), rec-FSH, rec-LH, and rec-FSH + rec-LH once a week. The results revealed that except for the SPE and the hCG groups, none of the groups exhibited a significant difference in GSI values. However, in vivo plasma E2 levels increased at the end of 4 weeks after rec-FSH treatment in female eels. Based on these results, it is suggested that rec-GTHs may have a positive effect on sexual maturation in female eels; however, further studies on complementary rec-protein production systems and additional glycosylation of rec-hormones are needed to elucidate hormone bioactivity in vivo and in vitro.

Potential Effect of the Intrusion of the Kuroshio Current into the South China Sea on Catches of Japanese Eel (Anguilla Japonica) in the South China Sea and Taiwan Strait

This study examined the effect of the intrusion of the Kuroshio Current (KC) into the South China Sea (SCS) and the Taiwan Strait (TS) (SCS–TS region) on changes in catches of larval A. japonica in the traditional fishing ground waters of Gaoping near southwestern Taiwan in the SCS–TS region. First, the oceanic environment and recruitment trends from 1967 to 2019 were investigated based on secondary data. Then, field surveys were conducted to obtain primary data regarding the intrusion of the KC into the SCS, as well as the changes in the fishing sites and catches of A. japonica in the fall and winter of 2014–2015. Hence, the association between oceanic conditions and the number of A. japonica migrating into the SCS–TS region was explored. From 1967 to 2019, the recruitment proportion in the fishing grounds that formed due to the Kuroshio Branch Current (PKSBC) fluctuated significantly. Overall, positive values were observed for the Oceanic Niño Index for each year with a PKSBC > 50%, corresponding to El Niño conditions. In each year with a PKSBC > 70%, a looping path and a warm–core eddy appeared.

Cell-Surface Loss of Constitutive Activating and Inactivating Mutants of Eel Luteinizing Hormone Receptors

The present study aimed to investigate the mechanism of cell surface receptor loss by two constitutively activating mutants (designated L469R, and D590Y) and two inactivating mutants (D417N and Y558F) of the luteinizing hormone receptor (LHR) in the Japanese eel Anguilla japonica, known to naturally occur in human LHR transmembrane domains. We investigated cell surface receptor loss using an enzyme-linked immunosorbent assay in HEK 293 cells. The expression level of wild-type eel LHR was considered to be 100%, and the expression levels of L469R and D417N were 97% and 101%, respectively, whereas the expression levels of D590Y and Y558F slightly increased to approximately 110% and 106%, respectively. The constitutively activating mutants L469R and D590Y exhibited a decrease in cell surface loss in a manner similar to that of wild-type eel LHR. The rates of loss of cell surface agonist-receptor complexes were observed to be very rapid (2.6-6.2 min) in both the wild-type eel LHR and activating mutants. However, cell surface receptor loss in the cells expressing inactivating mutants D417N and Y558F was slightly observed in the cells expressing inactivating mutants D417N and Y558F, despite treatment with a high concentration of agonist. These results provide important information on LHR function in fish and the regulation of mutations of highly conserved amino acids in glycoprotein hormone receptors.

Molecular characterization, expression patterns, and subcellular localization of a classical and a novel nonclassical MHC class I α molecules from Japanese eel Anguilla japonica

Major histocompatibility complex (MHC) molecules play a critical role in the immune response of vertebrate animals by presenting foreign antigens to T lymphocytes. In this study, we first cloned and identified a classical and a novel nonclassical MHC I α molecules from Japanese eel (Anguilla japonica), named as AjMHC I-UBA and AjMHC I-L, respectively. The full-length cDNA of AjMHC I-UBA contains an open reading frame (ORF) of 1047 bp encoding a predicted protein of 348 amino acids, while AjMHC I-L 1089 bp encodes 362 amino acids. The multiple alignment of the amino acid sequence showed that AjMHC I-UBA and AjMHC I-L consist of an N-terminal MHC I superfamily domain within α1 and α2 helices, an IgC-MHC I α3 domain, and a transmembrane region close to the C-terminal, which are similar to other fish and mammal species. Molecular polymorphism analysis showed that eight different major alleles of AjMHC I-UBA, named as AjMHC I-UBA*0101～1001, were identified from six Japanese eel individuals. Furthermore, a distinguishing signature of the nonclassical L-lineage genes specific motif “HINMTL”, including an N-glycosylation site (NXS/T), was present in the α3 domain of AjMHC I-L sequences. Although the predicted three-dimensional structures of AjMHC I-UBA and AjMHC I-L are similar to that of human MHC I α, phylogenetic analysis showed that these two protein molecules belong to classical MHC I UBA gene of U-lineage and nonclassical MHC I gene of L-lineage, respectively. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis revealed that the highest expression of AjMHC I-UBA and AjMHC I-L was found in the intestine and the expression level of AjMHC I-UBA in all of the examined tissues was significantly higher than that of AjMHC I-L. The expressions of AjMHC I-UBA and AjMHC I-L in liver, kidney and spleen were significantly induced following injection with the viral mimic poly I:C, LPS, and Aeromonas hydrophila infection. In vitro, the AjMHC I-UBA and AjMHC I-L transcripts of Japanese eel liver cells were significantly enhanced by the treatment of poly I:C or the stimulation of the high concentration of A. hydrophila. Subcellular localization showed that under natural conditions, AjMHC I-UBA and AjMHC I-L were uniformly distributed in the cytoplasm, and aggregated into spots or flakes in the cytoplasm after the stimulation of poly I:C or LPS. These results collectively suggested that AjMHC I-UBA and AjMHC I-L are important components possibly involved in Japanese eel defense against viral and bacterial infection. Taken together, these findings provide valuable insight into the immune function of MHC class I in the immune system of teleost.