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.

Preliminary insight into parental contributions to Japanese eel (Anguilla japonica) preleptocephali spawned on different nights.

Parentage sibship-inference analyses were conducted using mtDNA sequencing and six microsatellite genotypes of 182 Japanese eel preleptocephali that were collected from one net-tow near the West Mariana Ridge in May 2014. At least 328 parents were involved in producing the 182 preleptocephali, and several parents may have spawned a few times during 3 days of a spawning period. Half-sibs suggested that a few parents mated with 1-3 partners, indicating that the Japanese eel can form spawning aggregations in which several parents mate with each other in the ocean.

Distribution of anguillid leptocephali and possible spawning areas in the South Pacific Ocean.

Seven South Pacific anguillid eel species live from New Guinea to French Polynesia, but their spawning areas and life histories are mostly unknown despite previous sampling surveys. A July-October 2016 research cruise was conducted to study the spawning areas and times, and larval distributions of South Pacific anguillid eels, which included a short 155°E station-line northeast of New Guinea and five long transects (5-25°S, 160°E-140°W) crossing the South Equatorial (SEC) and other currents. This survey collected nearly 4000 anguilliform leptocephali at 179 stations using an Isaacs-Kidd Midwater Trawl accompanied by 104 CTD casts. Based on mor-phometric observations and DNA sequencing, 74 anguillid leptocephali were collected, which in the southern areas included 29 larvae of six species: Anguilla bicolor pacifica, A. marmorata, A. australis, A. reinhardtii, A. megastoma, and A. obscura (all anguillid species of the region were caught except A. dieffenbachii). Small A. australis (9.0-16.8 mm) and A. reinhardtii (12.4, 12.5 mm) leptocephali were collected south of the Solomon Islands, other A. australis (10.8-12.0 mm) larvae were caught northwest of Fiji along with an A. obscura (20.0 mm) larva, and an A. marmorata (7.8 mm) larva was collected near Samoa. Considering collection sites, larval ages from otolith analysis, and westward SEC drift, multiple spawning locations occurred from south of the Solomon Islands and the Fiji area (16-20 days old larvae) to near Samoa (19 days old larva) during June and July in areas where high-salinity Subtropical Underwater (STUW, ~150 m depth) and the warm, low-salinity surface Fresh Pool were present. Five long hydrographic sections showed the strong Fresh Pool in the west and the STUW formation area in the east.

Ghrelin and food acquisition in wild and cultured Japanese eel (Anguilla japonica).

To clarify the relationships between growth, endocrine status and habitat characteristics in Japanese eel (Anguilla japonica), plasma and stomach mRNA levels of ghrelin were examined in wild eels captured in the river and the bay, and in cultured eels during and after experimental fasting. Wild juvenile eels captured in freshwater habitats within the river showed significantly higher plasma and stomach mRNA levels of ghrelin than did fish obtained from brackish-water habitats within the bay. In cultured eels experimentally fasted for 4 weeks, plasma and stomach mRNA levels of ghrelin increased. After refeeding, the both parameters returned to the levels observed in continuously feeding control fish. In pigmented elvers, 2 months of feed restriction resulted in a significant increase in whole-body ghrelin mRNA. It is suggested that interaction between ghrelin and feeding is related to their habitats through differential food acquisition in fresh and brackish water environments.

Evaluation of downward movements of Japanese eel Anguilla japonica inhabiting brackish water areas.

This study evaluated the size and age distributions and otolith microchemistry of the Japanese eel Anguilla japonica in freshwater and brackish water areas in the Aki and Tsuchikawa rivers for 1 year, and in brackish water areas in the Asahi River for 3 years to understand the movements of Japanese eels between continental habitats of different salinity after recruitment (n = 759). For all three rivers, the total length (LT ) and age distributions were consistent; yellow eels captured in the upper brackish water (Aki River: 353.5 ± 77.4 mm and 3.0 ± 0.8 years; Tsuchikawa River: 287.7 ± 87.3 mm and 3.7 ± 1.3 years; Asahi River: 418.2 ± 112.1 mm and 4.2 ± 1.7 years) were smaller and younger than not only those in the fresh water of the two rivers but also those in the lowest brackish water sampling areas (Aki River: 436.0 ± 71.6 mm and 3.8 ± 1.1 years; Tsuchikawa River: 370.9 ± 121.7 mm and 4.9 ± 2.3 years; Asahi River: 558.5 ± 85.9 mm and 5.7 ± 1.7 years). In the Asahi River, these tendencies were found throughout the 3 years. Otolith analysis indicated that the majority of the eels captured in the lowest brackish water areas had moved down from upstream. These results suggest that Japanese eels inhabiting saline water generally move from the upper estuary as they grow. The upper estuary can be an important area for the management of this species because these eels spend their early continental growth life there.

Freshwater entry behaviour of a non-migratory stenohaline marine fish Takifugu snyderi.

We conducted salinity choice trials with the stenohaline marine species Takifugu snyderi to test their freshwater (FW) entry frequency in relation to starvation. The fish preferred to enter non-natal FW rather than remain in seawater. No relationship was detected between starvation and FW entry behaviour. Our results provide new empirical evidence of a stenohaline fish entering a non-natal osmotic environment. Further research on the entry of stenohaline species such as this one into lethal environments may help determine if this might help promote the evolution of diadromous life histories.

Morphological, ecological and physiological characteristics of downstream-migrating and non-migrating Pacific bicolor eels Anguilla bicolor pacifica.

This is the first study describing the morphological, ecological and physiological characteristics of two downstream-migrating and two non-migrating female Pacific bicolor eels, Anguilla bicolor pacifica. The total length and age of the downstream-migrating eels were 1005 mm and 10 years and 1110 mm and 11 years old, respectively, and those of the non-migrating eels were 892 mm and 8 years and 805 mm and 9 years, respectively. Silvering-related characteristics (silvering index, eye index, pectoral-fin index, gut-somatic index and swimbladder-somatic index) and reproductive physiological characteristics (gonado-somatic index, follicle diameter, oocyte stage, transcription of gonadotropins and concentration of sex steroids) of the migrating eels were more advanced than those of the non-migrating eels.

Reproductive physiological characteristics of tropical Celebes eels Anguilla celebesensis in relation to downstream migration and ovarian development.

Downstream-migrating (n = 64) and non-migrating (n = 21) female Celebes eels Anguilla celebesensis were captured from the Poso Lake-River system on Sulawesi Island, Indonesia, and their reproductive physiological characteristics were examined. A histological observation of the ovaries revealed that most non-migrating eels were at the perinucleolus (43%) or oil-droplet (48%) stage, whereas most migrating eels were at the early vitellogenic (36%) or midvitellogenic (61%) stage. Transcript levels of gonadotropin genes (fshb, lhb) in the pituitary gland and concentrations of sex steroids [11-ketotestosterone (11-KT), testosterone, 17ß-oestradiol (E2 )] in blood plasma of migrating eels were significantly higher than those of non-migrating eels. The fshb messenger (m)RNA levels were lower in perinucleolus and oil-droplet stages and then significantly increased in the early vitellogenic stage. The lhb mRNA levels in vitellogenic-stage eels were significantly higher than those in perinucleolus- and oil-droplet-stage eels. The 11-KT levels of eels at the oil-droplet and vitellogenic stages were significantly higher than those of eels at the perinucleolus stage. The E2 levels at the vitellogenic stage were significantly higher than those at the perinucleolus and oil-droplet stages. These dynamics of the reproductive hormones represented the physiological background of oogenesis in A. celebesensis that has remarkably well-developed oocytes just before downstream migration.

Spawning by the European eel across 2000 km of the Sargasso Sea.

It has been known for about a century that European eels have a unique life history that includes offshore spawning in the Sargasso Sea about 5000-7000 km away from their juvenile and adult habitats in Europe and northern Africa. Recently hatched eel larvae were historically collected during Danish, German and American surveys in specific areas in the southern Sargasso Sea. During a 31 day period of March and April 2014, Danish and German research ships sampled for European eel larvae along 15 alternating transects of stations across the Sargasso Sea. The collection of recently hatched eel larvae (≤12 mm) from 70° W and eastward to 50° W showed that the European eel had been spawning across a 2000 km wide region of the North Atlantic Ocean. Historical collections made from 1921 to 2007 showed that small larvae had also previously been collected in this wide longitudinal zone, showing that the spatial extent of spawning has not diminished in recent decades, irrespective of the dramatic decline in recruitment. The use of such a wide spawning area may be related to variations in the onset of the silver eel spawning migration, individual differences in their long-term swimming ability, or aspects of larval drift.

Changes in PRL Gene Expression During Upstream Movement of the Japanese Eel, Anguilla japonica.

Changes in mRNA levels of prolactin (PRL) during upstream movement were examined in juvenile Japanese eels, Anguilla japonica. Glass eels and elvers were collected from 2007 to 2009 near the entrance of Hamana Lake, and in a small inflowing stream, the Egawa River. Quantification of mRNA was performed by real-time PCR and expressed as whole-body content. PRL mRNA levels of glass eels caught in the coastal zone and tidal area were low. Eels that moved downward in the tidal zone and migrated upstream to enter into freshwater showed increased levels of PRL mRNA. These changes suggest the importance of up-regulation of PRL gene expression in juvenile eels during their upstream movement from seawater to fresh water, particularly in relation to hyperosmoregulation.

Mucosal galectin genes in all freshwater eels of the genus Anguilla.

In this study, we determined the genomic DNA sequences of the mucosal galectin-encoding genes from all 19 species and subspecies of the genus Anguilla. The nucleotide sequences of the galectin genes were c. 2.3-2.5 kb long and the organisation of their four exons and three introns was conserved in all species. An unusual sequence was found in the fourth exon of Anguilla reinhardtii, resulting in a unique deduced amino-acid sequence at the C-terminus. All six amino-acid residues important for ß-galactoside binding were conserved in three species, while one residue (R73 ) was substituted to K73 in the other 16 species-subspecies, including Anguilla marmorata. However, this substitution did not appear to affect the sugar-binding ability of galectins because the galectin of A. marmorata was previously shown to bind to lactose. We also discuss the molecular evolution of galectins among Anguilla spp. and the homologues previously identified in Conger myriaster.

Growth potential can affect timing of maturity in a long-lived semelparous fish.

Many diadromous fishes such as salmon and eels that move between freshwater and the ocean have evolved semelparous reproductive strategies, but both groups display considerable plasticity in characteristics. Factors such as population density and growth, predation risk or reproduction cost have been found to influence timing of maturation. We investigated the relationship between female size at maturity and individual growth trajectories of the long-lived semelparous European eel, Anguilla anguilla A Bayesian model was applied to 338 individual growth trajectories of maturing migration-stage female silver eels from France, Ireland, the Netherlands and Hungary. The results clearly showed that when growth rates declined, the onset of maturation was triggered, and the eels left their growth habitats and migrated to the spawning area. Therefore, female eels tended to attain larger body size when the growth conditions were good enough to risk spending extra time in their growth habitats. This flexible maturation strategy is likely related to the ability to use diverse habitats with widely ranging growth and survival potentials in the catadromous life-history across its wide species range.

Rapid genome reshaping by multiple-gene loss after whole-genome duplication in teleost fish suggested by mathematical modeling.

Whole-genome duplication (WGD) is believed to be a significant source of major evolutionary innovation. Redundant genes resulting from WGD are thought to be lost or acquire new functions. However, the rates of gene loss and thus temporal process of genome reshaping after WGD remain unclear. The WGD shared by all teleost fish, one-half of all jawed vertebrates, was more recent than the two ancient WGDs that occurred before the origin of jawed vertebrates, and thus lends itself to analysis of gene loss and genome reshaping. Using a newly developed orthology identification pipeline, we inferred the post-teleost-specific WGD evolutionary histories of 6,892 protein-coding genes from nine phylogenetically representative teleost genomes on a time-calibrated tree. We found that rapid gene loss did occur in the first 60 My, with a loss of more than 70-80% of duplicated genes, and produced similar genomic gene arrangements within teleosts in that relatively short time. Mathematical modeling suggests that rapid gene loss occurred mainly by events involving simultaneous loss of multiple genes. We found that the subsequent 250 My were characterized by slow and steady loss of individual genes. Our pipeline also identified about 1,100 shared single-copy genes that are inferred to have become singletons before the divergence of clupeocephalan teleosts. Therefore, our comparative genome analysis suggests that rapid gene loss just after the WGD reshaped teleost genomes before the major divergence, and provides a useful set of marker genes for future phylogenetic analysis.

Interspecific difference in downstream migratory season between two tropical eels, Anguilla celebesensis and Anguilla marmorata.

Downstream-migrating Anguilla celebesensis eels were predominant relative to Anguilla marmorata in October, November, December, January and February (75.9-92%), while no A. celebesensis occurred and A. marmorata were predominant in May and July (96-100%), at the outlet of Poso Lake, Sulawesi Island, Indonesia. Merging these results with those from published data suggests that most A. celebesensis start downstream migration during the early to middle rainy season, and A. marmorata migrate almost year-round with a peak from the late rainy to middle dry season.

Interspecific and sexual differences in riverine distribution of tropical eels Anguilla spp.

A total of 261 individuals of the four tropical eel species, Anguilla celebesensis, Anguilla marmorata, Anguilla bicolor pacifica and Anguilla interioris, were collected from 12 locations around Sulawesi Island, Indonesia, to gain knowledge about the riverine distribution of tropical eels. Anguilla marmorata was predominant in the lower reaches of Poso River (94·4% of total eel catch in the sampling area), Poso Lake (93·3%), three small inlet rivers of Tomini Bay (100%) and Laa River (92·3%). Anguilla celebesensis occurred frequently in the inlet rivers of Poso Lake (63·5%). Anguilla bicolor pacifica and Anguilla interioris were rare (1.5 and 0.4%, respectively). Otolith Sr:Ca ratio electron-probe micro analysis (EPMA) for individual migratory histories revealed that 15 A. celebesensis caught in Poso Lake and its inlet rivers were categorized into 14 river eels (Sr:Ca < 2·5) showing upstream migration seemingly at their elver stage and only one sea eel (Sr:Ca ≥ 6·0) that stayed in the marine habitat for the majority of its life after recruiting to Sulawesi Island before its late upstream migration. In A. marmorata, 19 examined eels from Poso Lake and its inlet rivers were all river eels, while 17 eels from the lower reaches of Poso River were two river eels, six sea eels and nine estuarine eels (2·5 ≤ Sr:Ca < 6·0) that mostly lived in the brackish water. The sex ratio of A. celebesensis was highly skewed towards a dominance of females (99%). In A. marmorata, females were predominant in Poso Lake (95·2%), its inlet rivers (94·7%) and Laa River (100%), while males were more frequent in the lower reaches of Poso River (76·5%) and small inlet rivers of Tomini Bay (94·1%). These results indicate that the riverine distribution pattern of tropical eels differs among species and between sexes.

A mechanism that maintains alternative life histories in a loggerhead sea turtle population.

Intrapopulation variation in habitat use is commonly seen among mobile animals, yet the mechanisms maintaining it have rarely been researched among untrackable species. To investigate how alternative life histories are maintained in a population of the loggerhead sea turtle (Caretta caretta), cumulative reproductive output was evaluated and compared between small planktivores inhabiting oceanic areas (with water depths > 200 m) and large benthivores inhabiting neritic areas (depths < 200 m) that sympatrically nested at Yakushima Island, Japan, from 1986 to 2011. In total, 362 nesting females sampled in three different years were classified into the two foraging groups based on stable isotope ratios in egg yolks. There were significant differences between the two foraging groups in most recorded life history parameters (clutch size, clutch frequency, breeding frequency, and remigration intervals), with the exception of emergence success. We did not find evidence of life history trade-offs, nor age-related changes in fecundity. Over the 26-year study period, we calculated a 2.4-fold greater reproductive output for neritic foragers than for oceanic ones, accounting for breeding and clutch frequency. Temporal consistencies in stable isotope ratios and remigration intervals within females suggested that female Japanese loggerheads show fidelity to respective foraging habitats throughout the adult stage. The large difference in productivity between the two groups was unlikely to be offset by the difference in survival during the period from aboveground emergence to first reproduction, suggesting that oceanic foragers have a lower level of fitness than neritic ones. Together with an absence of genetic structure between foraging groups, we infer that alternative life histories in a loggerhead turtle population are maintained by a conditional strategy.

A low trophic position of Japanese eel larvae indicates feeding on marine snow.

What eel larvae feed on in the surface layer of the ocean has remained mysterious. Gut contents and bulk nitrogen stable isotope studies suggested that these unusual larvae, called leptocephali, feed at a low level in the oceanic food web, whereas other types of evidence have suggested that small zooplankton are eaten. In this study, we determined the nitrogen isotopic composition of amino acids of both natural larvae and laboratory-reared larvae of the Japanese eel to estimate the trophic position (TP) of leptocephali. We observed a mean TP of 2.4 for natural leptocephali, which is consistent with feeding on particulate organic matter (POM) such as marine snow and discarded appendicularian houses containing bacteria, protozoans and other biological materials. The nitrogen isotope enrichment values of the reared larvae confirm that the primary food source of natural larvae is consistent only with POM. This shows that leptocephali feed on readily available particulate material originating from various sources closely linked to ocean primary production and that leptocephali are a previously unrecognized part of oceanic POM cycling.

Profiles of mRNA expression for prolactin, growth hormone, and somatolactin in Japanese eels, Anguilla japonica: The effect of salinity, silvering and seasonal change.

For understanding the functions of the growth hormone (GH)/prolactin (PRL)/somatolactin (SL) family of hormones, we examined pituitary mRNA expression of these hormones in anguillid eels in relation to salinity difference, silvering, and seasonal change. Female Japanese eels (Anguilla japonica) were collected in the brackish Hamana Lake and its freshwater rivers from July to December. To clarify the effect of salinity, the habitat use history of the eels were determined using otolith microchemistry. Expression levels of mRNA of each hormone were determined using real time PCR. Although GH and PRL have been known to be osmoregulatory hormones, there were no consistent differences in expression levels of these hormones between different salinity habitats. In contrast, SL mRNA expression was higher in eels from freshwater rivers than from the brackish lake. GH mRNA expression clearly decreased during silvering, whereas PRL and SL mRNA expression did not change. We also showed that PRL mRNA and SL mRNA decreased in the brackish lake and PRL mRNA increased in freshwater rivers from autumn to early winter. These findings provide basic knowledge for a further understanding of the role of these hormones.

Migration Behavior of Anguilla celebesensis Silver Eels within their Tomini Bay Spawning Area.

The tropical Celebes eel, Anguilla celebesensis, has a short migration between its spawning and growth habitats. Its spawning areas were hypothesized to be in Tomini Bay and the Celebes Sea after collecting their small leptocephali. However, there is no information about the silver eel oceanic spawning migration behavior of A. celebesensis. To better understand their short-distance spawning migration behavior, four large female silver eels (Eel 1-4) were equipped with pop-up satellite archival tags (PSATs) and released near the mouth of the Poso River in Tomini Bay of Sulawesi Island on 22 February (Eel 1-3) and 11 March 2010 (Eel 4). All PSATs ascended in Tomini Bay and transmitted their data. Eel 3 and 4 provided clear records of consistent diel vertical migration (DVM: eight days-Eel 3, 13 days-Eel 4) with daytime dives to mean depths of 444.7 m (Eel 3) and 539.0 m (Eel 4), where mean temperatures were 9.1°C (Eel 3) and 7.7°C (Eel 4), and nighttime ascents to mean depths of 132.8 m (Eel 3) and 112.4 m (Eel 4), where mean temperatures were 20.6°C (Eel 3) and 23.4°C (Eel 4). Eel 3 and 4 started to dive to deeper water around nautical dawn and swam up to shallower water around sunset. During nighttime, both eels swam in deeper and colder water during nights with moonlight than during nights without moonlight, and there was a negative linear relationship between experienced water temperatures with the moon in the sky and the lunar age for the eels. The A. celebesensis daily rhythm of DVM behaviors was similar to spawning-migration DVM behaviors of other anguillid species. Essential life history characteristics of A. celebesensis appear to be a short migration between freshwater growth habitat and ocean spawning habitat, and high GSI values with advanced gonadal development in downstream-migrating silver eels.

Oceanic biology: spawning of eels near a seamount.

Discoveries of the larvae of the European and American eels, Anguilla anguilla and A. rostrata, in the Sargasso Sea and of the Japanese eel, A. japonica, in the Philippine Sea indicate that these freshwater eels migrate thousands of kilometres into the open ocean to spawn. Here we pinpoint a spawning location for Japanese eels after genetically identifying newly hatched larvae that we collected from the site. The restricted size of this spawning area ensures that the eel larvae enter a particular current that transports them to the freshwater areas in east Asia where they mature, and it also prevents them from being carried southwards away from their species range by a different local current.