Recent insights into egg quality and larval vitality of the European eel Anguilla anguilla.

To date, the eel industry still depends on wild-caught juveniles that are grown to marketable size. There is an urgent need to close the eel life cycle in captivity to make aquaculture independent of the natural population. With this artificial reproduction protocol, yolk-sac larvae can be produced but egg quality may be impaired. Low survival rates and high deformity rates are frequently observed during the first week after hatching. Over the past four years, we have conducted studies with the aim to optimize the artificial reproduction protocol, thereby focussing on increasing egg and larval quality. Weekly carp or salmon pituitary extract (PE) treatment was successfully replaced with recombinant gonadotropins (rGTHs) to mature female eels and produce larvae. 17α,20ß-dihydroxy-4-pregnen-3-one (DHP) was replaced with upstream precursor progesterone (P) to induce the endogenous production of DHP by the female eel. DHP and P were found equally potent in inducing oocyte maturation and ovulation. The effects of antibiotics on larval survival and the occurrence of deformities were investigated. Antibiotic treatment increased survival and decreased the occurrence of deformities indicating bacterial infection as an important cause. A deformity determination key for young eel larvae has been developed that provides a framework of reference for larval deformities which will be instrumental with gaining insights on the reasons behind each larval deformity. These improvements of the artificial reproduction protocol and hatchery practices will contribute to the production of robust eel larvae that survive, grow and metamorphose into juveniles that will later be able to reproduce in captivity.

Superoxidase dismutases (SODs) in the European eel: Gene characterization, expression response to temperature combined with hormonal maturation and possible migratory implications.

Superoxide dismutases (SODs) are antioxidant enzymes that protect cells from oxidation. Three SODs have been identified in mammals, but there is limited information in teleosts. This study investigates SODs in the European eel and their expression patterns during testis maturation. Phylogenetic and synteny analyses revealed SODs paralogs and their evolution in vertebrates. The eel possesses one SOD1 and two SOD2/3 (a and b), indicating SOD2 and SOD3 duplication in elopomorphs. SODs expression were then evaluated in various male and female tissues. SOD1 is more expressed in females, while SOD2a and SOD2b dominate brain-pituitary-gonad tissues in both sexes. SOD3a showed predominant expression in the ovary and the male livers, whereas SOD3b was found in the pituitary and brain of both sexes. The effects of different maturation protocols (standard hormonal treatment vs. same protocol preceded with cold seawater pre-treatment) on SODs expression during testis maturation were evaluated. Salinity increase at the onset of standard treatment at 20 °C, simulating early migration, upregulated SOD1, SOD2a, and SOD2b, coinciding with spermatogonia type A differentiated cells dominance. Thereafter, SOD2a and SOD3a decreased, while SOD2b increased during hormonal treatment-induced spermatogenesis. Pre-treatment with seawater at 10 °C, mimicking the conditions at the beginning of the seawater migration, downregulated SOD1 but increased SOD3a expression. Finally, the standard hormonal treatment, replicating spawning at higher temperatures, downregulated SOD1 in eels without any pre-treatment while SOD2a expression increased in pre-treated eels. This study revealed tissue-specific, sex-dependent, and maturation-related SOD expression patterns, predicting SODs dynamic expression profiles during their reproductive migration.

Unravelling the life-history patterns and habitat preferences of the Japanese eel (Anguilla japonica) in the Pearl River, China.

Eels have fascinated biologists for centuries due to their amazing long-distance migrations between freshwater habitats and very distant ocean spawning areas. The migratory life histories of the Japanese eel, Anguilla japonica, in the waters of south China are not very clear despite its ecological importance, and the need for fishery regulation and management. In this study, strontium (Sr) and calcium (Ca) microchemical profiles of the otoliths of silver eels were measured by X-ray electron probe microanalysis based on data collected from different habitats (including freshwater and brackish habitats), in the large subtropical Pearl River. The corresponding habitat preference characteristics were further analysed using redundancy analysis (RDA). A total of 195 Japanese eels were collected over 6 years. The collected individuals ranged from 180 to 771 mm in total length and from 8 to 612 g in body weight. Two-dimensional pictures of the Sr:Ca concentrations in otoliths revealed that the A. japonica in the Pearl River are almost entirely river eels, spending the majority of their lives in fresh water without exposure to salt water, while the catadromous migration time has delayed about 1 month in the Pearl River estuary in the past 20 years. RDA analysis further indicated that juveniles and adults preferred water with high salinity and high tide levels. Youth preferred habitats with high river fractals. Our findings contribute to a growing body of evidence showing that the eels are extremely scarce currently and conservation measures against them are imminent, including the protection of brackish and freshwater areas where they live in south China.

Wastewater plumes can act as non-physical barriers for migrating silver eel.

Non-physical barriers for migrating fish, such as effluent plumes discharged by sewage treatment plants (WWTPs), are hardly considered, and field studies on this topic are very scarce. The encounter with these plumes however may evoke behavioural responses in fish and could delay or (partially) block the migration. In this study, the behavioural responses of 40 acoustically-tagged silver eel (Anguilla anguilla) were monitored in situ, when confronting a WWTP effluent plume during their downstream migration in the canal Eems, the Netherlands. Their behavioural responses and the potential blocking effect of the plume were assessed using a 2D and 3D telemetry design displayed in the waterway, and matched to a modelled and calibrated WWTP effluent plume. When confronted with the WWTP effluent plume during their downstream migration, 22 of the silver eels (59%) showed an avoidance response, varying from lateral diverting to multiple turning in the vicinity of the effluent plume. Nineteen out of these 22 (86%) eventually passed the study site. No silver eel showed attraction to the plume. Delays in migration were from several hours up to several days. Due to the strong variation in discharged volumes and flow velocity of the receiving canal, the WWTP plume did not always flow over the full width of the canal. As a result, numerous migratory windows, where silver eels could pass the WWTP while avoiding direct contact with the plume, remained available in time. When discharge points cannot be avoided, reduced or restricted to areas that are not preferred as fish migration routes, discharge points should be designed such, that the chance is limited that a waterway is (temporarily) impacted over its full width.

Exploring first-feeding diets for European eel larval culture: Insights at morphological, nutritional, and molecular levels.

Closing the life cycle of European eel (Anguilla anguilla) in captivity is targeted to provide a sustainable, year-round supply of juveniles for aquaculture. Present focus is on the nutritional requirements during the larval first-feeding period. In this study, three experimental diets were tested on hatchery-produced European eel larvae from the onset of the first-feeding stage commencing 10 days post hatch (dph) until 28 dph. Larval mortality was recorded daily, while sampling was conducted at regular intervals to record larval biometrics and analyze the expression of genes related to digestion, appetite, feed intake and growth. Two periods of high mortality were identified: the first appeared shortly after introduction of feeds (10-12 dph), while the second occurred 20-24 dph, indicating the "point of no return". This interpretation was supported at the molecular level by the expression of the gene encoding the "hunger hormone" ghrelin (ghrl) that peaked at 22 dph in all dietary trials, suggesting that most larvae were fasting. However, in larvae fed diet 3, ghrl expression was downregulated beyond 22 dph, which indicated that those larvae were no longer starving at this stage, while upregulation of genes encoding the major digestive enzymes (try, tgl, and amyl2a) advocated their healthy development. Moreover, for larvae fed diet 3, the expression of those genes as well as genes for feed intake (pomca) and growth (gh) continued to increase towards 28 dph. These results together with the registered highest survival, largest dry weight increase, and enhanced biometrics (length and body area) pointed to diet 3 as the best-performing. As a whole, this first-feeding study represents a landmark being the first to document European eel larval growth and survival beyond the point of no return, providing novel insights into the molecular development of digestive functions during the first feeding stage.

Effects of preservation by ethanol on δ13 C and δ15 N of three tissues of the critically endangered European eel Anguilla anguilla.

The temporal effects of ethanol preservation on the δ13 C and δ15 N values of tissues excised from European eel Anguilla anguilla were assessed. Preservation significantly enriched 13 C values of fin and mucus but not dorsal muscle. The 13 C enrichment occurred in the initial 15 days of preservation and was independent of initial eel mass. Tissue preservation effects on δ15 N values were negligible. These tissue-specific isotopic shifts should be considered when ethanol-preserved eel samples are used.

Water temperature and precipitation stimulate small-sized Japanese eels to climb a low-height vertical weir.

Although river-crossing structures can have a detrimental effect on the movement and distribution of anguillid eels (genus Anguilla), they have inhabited locations upstream of river-crossing structures in many rivers. However, the growth stage in which the eels start to climb river-crossing structures remains unclear. In this study, we directly observed, using infrared video camera systems, that the Japanese eel (Anguilla japonica) climbs a low-height vertical weir as a migratory barrier, confirming the ability of eels to climb over a low-height weir within a river. All eels climbed the weir at night, between sunset and sunrise. We observed that the total length of Japanese eels that climbed the weir ranged from 60 to 140 mm, suggesting that eels inhabiting the upstream area of a weir climbed it when they were small and then lived and grew upstream. Moreover, the general additive model showed considerable effects of water temperature and precipitation on eel climbing, suggesting that water temperature and precipitation are important for its activation. The results of this study also show the effectiveness of infrared video cameras in monitoring eel climbing, which could be applied to monitor fish migratory behavior through fish passages. The findings of this study are useful for the comprehensive management and conservation of wild eel stocks.

Glass eel (Anguilla anguilla L. 1758) feeding behaviour during upstream migration in an artificial waterway.

The transition from marine to fresh water is a challenging task for juvenile eels. This critical step in the early eels' life is preceded by a metamorphosis from the oceanic larval to the continental glass eel stage, requiring major energy-demanding morphological, physiological and behavioural modifications during which time these animals do not feed. The success of the glass eels' inland migration after metamorphosis will largely depend on remaining energy levels, which can be supplemented only by resuming food uptake. Although it is crucial for their survival and the maintenance of the population, the feeding behaviour of glass eels is still an understudied aspect of the eels' complex life cycle. Many uncertainties about the phenology, diet, potential prey preferences and their relation with migration modus (migratory vs. sedentary) still remain. In this study, the authors analysed the stomach and gut contents of 458 European glass eels (Anguilla anguilla L. 1758) captured in a drainage canal connecting a small mesotidal estuary with an adjacent polder area during the spring migration seasons of 2016 and 2017. They demonstrated that although glass eels started feeding briefly upon arrival in the estuary, food uptake for early arrivals was restricted to a minority that sparsely feed on detritus and some worm-like benthic invertebrates. Along the season, food uptake intensified eventually engaging all glass eels and their dietary palette diversified including a wide array of planktonic and benthic organisms. Crustacean plankton (mainly cyclopoid copepods) was an important part of the glass eel diet, whereas benthic oligochaetes were less abundant as food source in spite of their high presence in the sediments. No clear differences in feeding behaviour could be observed between migratory and sedentary glass eels. This study showed that glass eels can use highly artificial and dynamic drainage canals as feeding ground during their critical marine/freshwater transition. This outcome is also a plea to improve the accessibility of alternative (unnatural) migration routes between the ocean and suitable freshwater growth habitats for the European eel.

Impact of nickel mining in New Caledonia on marbled eels Anguilla marmorata.

New Caledonia is particularly affected by nickel open pit mining activities because of the presence of ultramafic soils rich in metals. The particles dispersed by atmospheric transport and soil erosion during the excavation of nickel end up by deposition or leaching in rivers where they may be bioaccumulated by organisms living downstream the mines. Despite alarming freshwater metals concentrations, no study investigated the level of their bioaccumulation in eels, and if high bioaccumulation levels occur, the potential consequences on their health. The aim of this study was to determine how eels Anguilla marmorata are impacted in situ by metals issued from mining activity by measuring: morphometric parameters; metal concentrations in tissues and organs and transcription levels of target genes encoding proteins involved in several metabolic key functions. Among organs, liver was found to be the most affected by mining with average nickel concentrations of 5.14 mg/kg versus 1.63 mg/kg for eels away from mines leading to dysregulation of numerous genes involved in oxidative stress, DNA repair, apoptosis, reproduction and both lipid and mitochondrial metabolisms. This study should allow us to define in an integrated way if metals released by mining activities influence metals bioaccumulation in eels and induce biological effects.

Response of upstream migrating juvenile European eel (Anguilla anguilla) to electric fields: Application of the marginal gains concept to fish screening.

The decline in European eel (Anguilla anguilla) recruitment over the past half-century is partly due to river infrastructure that delays or blocks upstream migration to rearing habitat. Stimuli, such as electricity, can be used to modify the behaviour of downstream moving fish and guide them to preferred routes of passage at river infrastructure; but research on upstream migrating juvenile eel remains limited. The response of upstream migrating juvenile eel exposed to pulsed direct current (PDC) electric fields was investigated using a recirculatory flume. Eel were presented a choice of two routes upstream under either: (1) a treatment condition, in which the selection of one route resulted in exposure to High Electric Field (HEF) strength that was between 1.5-2 times stronger than the Low Electric Field (LEF) strength encountered in the alternative route; or (2) a control in which the electric field was absent in both routes. Under the treatment, five different mean HEF strengths (0.53, 0.77, 1.22, 2.17 and 3.74 Vcm-1) were tested at one of two frequencies (2 and 10 Hz). Route choice, distance downstream of the first set of electrodes at which an initial response was observed and avoidance behaviours (acceleration, retraction, switching and rejection) were compared among treatments. For the 1.22, 2.17 and 3.74 Vcm-1 and under 2 Hz, eel preferred to pass the LEF route. Avoidance was greater in the HEF route and positively related to field strength. The distance of the initial response did not differ between routes, field strengths or frequency. Upstream migrating eel avoided electric fields indicating potential to develop this approach for fish guidance. Further work is needed to test prototypes in field settings, particularly in combination with traditional physical screens to water intakes as part of a process of applying the concept of marginal gains to advance environmental impact mitigation technology.

Metabolism of hyaluronic acid during early development of the Japanese eel, Anguilla japonica.

Hyaluronic acids (hyaluronans, HAs) are glycosaminoglycans produced in the bodies of Anguilliforme and Elopiforme leptocephali, and HA is thought to serve as a metabolic energy source during planktonic life-stages. To examine this hypothesis, we investigated the dynamics of HA during early growth of the Japanese eel (Anguilla japonica), including during metamorphosis. From histochemical observations in the fully grown leptocephalus, HA occupied approximately 40-50% of the cross-sectional area and muscle tissue occupied less than 20%. However, the HA and water content are at a maximum during leptocephalus, decreasing during metamorphosis. We found that during leptocephalus, HA is actively accumulated in the body and plays a role in specific density adjustment, facilitating planktonic life. It was thought that after metamorphosis the role of HA in facilitating floating would end, and it would be metabolized to glucose and/or triglyceride.

Non-lethal sampling for the stable isotope analysis of the critically endangered European eel Anguilla anguilla: how fin and mucus compare to dorsal muscle.

Ecological studies on the critically endangered European eel Anguilla anguilla often incorporate stable isotope analysis that typically uses dorsal muscle sampled from euthanised eels. To minimise the lethal sampling of imperilled populations, fin tissue and/or epidermal mucus can provide non-lethal alternatives to muscle. The results here indicate that δ13 C and δ15 N values of both eel fin and mucus are not significantly different from those of muscle and can be applied directly in comparative SI studies.

Mapping silver eel migration routes in the North Sea.

Recent developments in tracking technology resulted in the mapping of various marine spawning migration routes of the European eel (Anguilla anguilla). However, migration routes in the North Sea have rarely been studied, despite many large European rivers and hence potential eel growing habitat discharge into the North Sea. In this study, we present the most comprehensive map to date with migration routes by silver European eels in the North Sea and document for the first time successful eel migration through the English Channel. Migration tracks were reconstructed for 42 eels tagged in Belgium and 12 in Germany. Additionally, some eels moved up north to exit the North Sea over the British Isles, confirming the existence of two different routes, even for eels exiting from a single river catchment. Furthermore, we observed a wide range in migration speeds (6.8-45.2 km day-1). We hypothesize that these are likely attributed to water currents, with eels migrating through the English Channel being significantly faster than eels migrating northward.

Impairments in spatial learning by telencephalic lesions in Japanese eels (Anguilla japonica).

This study aimed to use Japanese eels (Anguilla japonica) as subjects to examine the effects of telencephalic lesions on spatial learning. Ten Japanese eels were trained on a Morris-type spatial learning task. Four pipes were placed in a pool; however, the eels could hide in only one of these pipes. The learning task ensured that the eels learned about the position of the open pipe. Subsequently, their telencephalons were damaged. The lesioned eels could not maintain their learning and demonstrated deficits in re-learning as some of them were unable to relearn the task. An analysis of the lesion sizes revealed that while damage to the dorsolateral pallium correlates with maintenance of learning, damage to the dorsomedial pallium correlates with re-learning.

A novel hyperbaric swimming respirometer allows the simulation of varying swimming depths in fish respirometry studies.

The understanding of swimming physiology and knowledge on the metabolic costs of swimming are important for assessing effects of environmental factors on migratory behavior. Swim tunnels are the most common experimental setups for measuring swimming performance and oxygen uptake rates in fishes; however, few can realistically simulate depth and the changes in hydrostatic pressure that many fishes experience, e.g. during diel vertical migrations. Here, we present a new hyperbaric swimming respirometer (HSR) that can simulate depths of up to 80 m. The system consists of three separate, identical swimming tunnels, each with a volume of 205 L, a control board and a storage tank with water treatment. The swimming chamber of each tunnel has a length of 1.40 m and a diameter of 20 cm. The HSR uses the principle of intermittent-flow respirometry and has here been tested with female European eels (Anguilla anguilla). Various pressure, temperature and flow velocity profiles can be programmed, and the effect on metabolic activity and oxygen consumption can be assessed. Thus, the HSR provides opportunities to study the physiology of fish during swimming in a simulated depth range that corresponds to many inland, coastal and shelf waters.

Characterization of triglycerides during early development of the Japanese eel (Anguilla japonica).

Japanese eels store lipids in the peritoneal cavity at the glass eel stage. These lipids are presumed to be consumed as an energy source during migration from oceanic metamorphosing sites to coastal Japan. In this study we investigated the distribution and amount of triglyceride in vivo using artificially bred Japanese eel larvae. Triglycerides accumulated in vivo from the early larval stage onward, until they reached a peak volume in fully grown leptocephali, and subsequently gradually decreased during metamorphosis. Furthermore, during the late metamorphic stages of the artificially bred glass eel, triglycerides were stored mainly in the peritoneal cavity, as in wild glass eels. These observations point to a strategy that the Japanese eels use in consuming the triglycerides derived from leptocephali during metamorphosis (when they do not feed) as a source of energy.

First tracking of the oceanic spawning migrations of Australasian short-finned eels (Anguilla australis).

Anguillid eel populations have declined dramatically over the last 50 years in many regions of the world, and numerous species are now under threat. A critical life-history phase is migration from freshwater to distant oceans, culminating in a single life-time spawning event. For many anguillids, especially those in the southern hemisphere, mystery still shrouds their oceanic spawning migrations. We investigated the oceanic spawning migrations of the Australasian short-finned eel (Anguilla australis) using pop-up satellite archival tags. Eels were collected from river estuaries (38° S, 142° E) in south-eastern temperate Australia. In 2019, 16 eels were tracked for up to about 5 months, ~ 2620 km from release, and as far north as the tropical Coral Sea (22° S, 155° E) off the north-east coast of Australia. Eels from southern Australia appeared to access deep water off the Australian coast via two main routes: (i) directly east via Bass Strait, or (ii) south-east around Tasmania, which is the shortest route to deep water. Tagged eels exhibited strong diel vertical migrations, alternating between the warm euphotic zone (~ 100-300 m, 15-20 °C) at night and the mesopelagic zone (~ 700-900 m, 6-8 °C) during the day. Marine predators, probably lamnid sharks, tuna, or marine mammals, ended many eel migrations (at least ~ 30%), largely before the eels had left the Australian continental shelf. The long and risky marine migrations of Australasian eels highlight the need for better information on the processes contributing to eel mortality throughout the life cycle, including the impacts of future changes to oceanic currents, predator abundance and direct anthropogenic disturbances.

Feeding habitat and silvering stage affect lipid content and fatty acid composition of European eel Anguilla anguilla tissues.

Lipids, particularly fatty acids (FAs), are major sources of energy and nutrients in aquatic ecosystems and play key roles during vertebrate development. The European eel Anguilla anguilla goes through major biochemical and physiological changes throughout its lifecycle as it inhabits sea- (SW), and/or brackish- (BW) and/or freshwater (FW) habitats. With the ultimate goal being to understand the reasons for eels adopting a certain life history strategy (FW or SW residency vs. 'habitat shifting'), we explored differences in lipid content and FA composition of muscle, liver and eyes from eels collected across Norwegian SW, BW and FW habitats, and at different lifecycle stages (yellow to silver). FW and SW eels had a higher lipid content overall compared to BW eels, reflecting differences in food availability and life history strategies. SW eels had higher proportions of certain monounsaturated FAs (MUFAs; 18:1n-9, 20:1n-9), and of the essential polyunsaturated FAs 20:5n-3 (eicosapentaenoic acid, EPA) and 22:6n-3 (docosahexaenoic acid) than FW eels, reflecting a marine-based diet. In contrast, the muscle of FW eels had higher proportions of 18:3n-3, 18:2n-6 and 20:4n-6 (arachidonic acid), as is typical of FW organisms. MUFA proportions increased in later stage eels, consistent with the hypothesis that the eels accumulate energy stores prior to migration. In addition, the decrease of EPA with advancing stage may be associated with the critical role that this FA plays in eel sexual development. Lipid and FA information provided further understanding of the habitat use and overall ecology of this critically endangered species.

Environmental salinity influences the branchial expression of TCR pathway related genes based on transcriptome of a catadromous fish.

Environmental salinity not only affects the physiological processes such as osmoregulation and hormonal control, but also changes the immune system in fishes. Studies are limited in fish on the roles of the T cell receptor (TCR)-related genes in relation to changes in environmental salinity. A large group of salinity-challenged transcripts was obtained in gills of marbled eel (Anguilla marmorata). Moreover, bioinformatic ways were used to identify the enriched TCR pathway related genes which were significantly different expressed in fresh water (FW), brackish water (BW) and seawater (SW). Meanwhile, the RT-qPCR results were validated and consistent with the RNA-seq results. TCR a, TCR b, CD45, CD28, PI3K, LCK and LAT were up-regulated when the salinity increases in BW and SW, which connected with the related signaling pathways (Ras-MAPK and PKC pathway). CD4 and Zap70 were down-regulated when the salinity increases in BW and SW, which connected with the PLC pathway. The research offers a novel viewpoint to explore the immune pathways including the TCR pathway in fish based on transcriptome.

Impact of downstream passage through hydropower plants on the physiological and health status of a critically endangered species: The European eel Anguilla anguilla.

Hydropower plants (HPPs) are a source of "green" energy but also a threat to migrating fish such as the European eel (Anguilla anguilla) owing to the disruption of river connectivity and the obstruction of downstream migration. The impact of HPP are well-documented in terms of fish survival and damages but there is no available information concerning the condition of surviving and unharmed fish. The aim of this study is to assess the impact of the passage through HPP on the survival, the physiological and health status of adult eels. Two trials were carried with variants of the Kaplan turbine - one of the most common types in Europe. After a deliberate passage through the turbines, we studied direct mortality, external and internal damages, stress and immune biomarkers such as plasma cortisol and glucose levels, alternative complement (ACH50), lysozyme and peroxidase activities, and total immunoglobulin (Ig) content. Our results showed a lower survival and a higher external and internal damages rates in the HPP groups. Glucose levels, ACH50, lysozyme and peroxidase activities and TIgc were also affected by the passage depending on HPP characteristics. Those findings suggest a greater energy expenditure and a disruption in innate immunity due to this passage. HPPs can not only have an impact in terms of direct mortality and injuries but also affect the physiological and health condition of the surviving eels. This impact may explain the delayed mortality observed in telemetric studies and the passage through many HPPs may compromise the ability of adult eels to migrate successfully to the ocean.