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Fish embryos may be vulnerable to seawater acidification resulting from anthropogenic carbon dioxide (CO2) emissions or from excessive biological CO2 production in aquaculture systems. This study investigated CO2 effects on embryos of the European eel (Anguilla anguilla), a catadromous fish that is considered at risk from climate change and that is targeted for hatchery production to sustain aquaculture of the species. Eel embryos were reared in three independent recirculation systems with different pH/CO2 levels representing "control" (pH 8.1, 300 μatm CO2), end-of-century climate change ("intermediate", pH 7.6, 900 μatm CO2) and "extreme" aquaculture conditions (pH 7.1, 3000 μatm CO2). Sensitivity analyses were conducted at 4, 24, and 48 hours post-fertilization (hpf) by focusing on development, survival, and expression of genes related to acute stress response (crhr1, crfr2), stress/repair response (hsp70, hsp90), water and solute transport (aqp1, aqp3), acid-base regulation (nkcc1a, ncc, car15), and inhibitory neurotransmission (GABAAα6b, Gabra1). Results revealed that embryos developing at intermediate pH showed similar survival rates to the control, but egg swelling was impaired, resulting in a reduction in egg size with decreasing pH. Embryos exposed to extreme pH had 0.6-fold decrease in survival at 24 hpf and a 0.3-fold change at 48 compared to the control. These observed effects of acidification were not reflected by changes in expression of any of the here studied genes. On the contrary, differential expression was observed along embryonic development independent of treatment, indicating that the underlying regulating systems are under development and that embryos are limited in their ability to regulate molecular responses to acidification. In conclusion, exposure to predicted end-of-century ocean pCO2 conditions may affect normal development of this species in nature during sensitive early life history stages with limited physiological response capacities, while extreme acidification will negatively influence embryonic survival and development under hatchery conditions., Competing Interests: The authors have declared that no competing interests exist.

To meet the growing demand of fish feed for aquaculture, an increasing proportion of marine ingredients are being replaced by blends of plant products. However, the total replacement of marine ingredients in salmonid diets impairs fish performance. This is particularly true during the early fry stage and this stage is therefore considered of particular importance. In rainbow trout (RBT), the existence of a genetic variability to survive and grow with plant-based diets devoid of marine ingredients has now been proved, but the mechanisms behind are little studied especially at early stage. To investigate these, we analysed the whole transcriptome of three isogenic lines of RBT fry, which have similar growth when fed a marine resources-based diet (M diet) but which highly differ in their responses to a plant-based diet (V diet). Analysis of transcriptomes profiles revealed 1740, 1834 and 246 probes differentially expressed among the three genotypes when fed the V diet. The use of these lines led to the discovery of potential molecular markers linked to plant-based diet utilisation, some of them belonging to new pathways, never described before. An important number of genes was related to immunity, but further investigations are needed to better understand the difference between the genotypes in their immune status response to V diet exposure. Finally, differences in expression of genes related to feed intake and sensory perception among genotypes suggested that the mechanisms underlying the differences in growth on plant-based diet are closely linked to diet acceptance. Research on plants components affecting feed intake should be thus further explored., Competing Interests: The authors have declared that no competing interests exist.

European eel (Anguilla anguilla) is a euryhaline species, that has adapted to cope with both, hyper- and hypo-osmotic environments. This study investigates the effect of salinity, from a morphological and molecular point of view on European eel larvae reared from 0 to 12 days post hatch (dph). Offspring reared in 36 practical salinity units (psu; control), were compared with larvae reared in six scenarios, where salinity was decreased on 0 or 3 dph and in rates of 1, 2 or 4 psu/day, towards iso-osmotic conditions. Results showed that several genes relating to osmoregulation (nkcc2α, nkcc2β, aqp1dup, aqpe), stress response (hsp70, hsp90), and thyroid metabolism (thrαA, thrαB, thrβB, dio1, dio2, dio3) were differentially expressed throughout larval development, while nkcc1α, nkcc2β, aqp3, aqp1dup, aqpe, hsp90, thrαA and dio3 showed lower expression in response to the salinity reduction. Moreover, larvae were able to keep energy metabolism related gene expression (atp6, cox1) at stable levels, irrespective of the salinity reduction. As such, when reducing salinity, an energy surplus associated to reduced osmoregulation demands and stress (lower nkcc, aqp and hsp expression), likely facilitated the observed increased survival, improved biometry and enhanced growth efficiency. Additionally, the salinity reduction decreased the amount of severe deformities such as spinal curvature and emaciation but also induced an edematous state of the larval heart, resulting in the most balanced mortality/deformity ratio when salinity was decreased on 3 dph and at 2 psu/day. However, the persistency of the pericardial edema and if or how it represents an obstacle in further larval development needs to be further clarified. In conclusion, this study clearly showed that salinity reduction regimes towards iso-osmotic conditions facilitated the European eel pre-leptocephalus development and revealed the existence of highly sensitive and regulated osmoregulation processes at such early life stage of this species., Competing Interests: The authors have declared that no competing interests exist.

Temperature is important for optimization of rearing conditions in aquaculture, especially during the critical early life history stages of fish. Here, we experimentally investigated the impact of temperature (16, 18, 20, 22 and 24°C) on thermally induced phenotypic variability, from larval hatch to first-feeding, and the linked expression of targeted genes [heat shock proteins (hsp), growth hormone (gh) and insulin-like growth factors (igf)] associated to larval performance of European eel, Anguilla anguilla. Temperature effects on larval morphology and gene expression were investigated throughout early larval development (in real time from 0 to 18 days post hatch) and at specific developmental stages (hatch, jaw/teeth formation, and first-feeding). Results showed that hatch success, yolk utilization efficiency, survival, deformities, yolk utilization, and growth rates were all significantly affected by temperature. In real time, increasing temperature from 16 to 22°C accelerated larval development, while larval gene expression patterns (hsp70, hsp90, gh and igf-1) were delayed at cold temperatures (16°C) or accelerated at warm temperatures (20-22°C). All targeted genes (hsp70, hsp90, gh, igf-1, igf-2a, igf-2b) were differentially expressed during larval development. Moreover, expression of gh was highest at 16°C during the jaw/teeth formation, and the first-feeding developmental stages, while expression of hsp90 was highest at 22°C, suggesting thermal stress. Furthermore, 24°C was shown to be deleterious (resulting in 100% mortality), while 16°C and 22°C (~50 and 90% deformities respectively) represent the lower and upper thermal tolerance limits. In conclusion, the high survival, lowest incidence of deformities at hatch, high yolk utilization efficiency, high gh and low hsp expression, suggest 18°C as the optimal temperature for offspring of European eel. Furthermore, our results suggest that the still enigmatic early life history stages of European eel may inhabit the deeper layer of the Sargasso Sea and indicate vulnerability of this critically endangered species to increasing ocean temperature.

The objective of this study was to evaluate the combined effects of thermal acclimation and n-3 highly unsaturated fatty acids (n-3 HUFA) content of the food source on the aerobic capacities of fish in a thermal changing environment. The model used was the golden grey mullet Liza aurata, a species of high ecological importance in temperate coastal areas. For four months, fish were exposed to two food sources with contrasting n-3 HUFA contents (4.8% ecosapentaenoic acid EPA + docosahexaenoic acid DHA on the dry matter DM basis vs. 0.2% EPA+DHA on DM) combined with two acclimation temperatures (12°C vs. 20°C). The four experimental conditions were LH12, LH20, HH12 and HH20. Each group was then submitted to a thermal challenge consisting of successive exposures to five temperatures (9°C, 12°C, 16°C, 20°C, 24°C). At each temperature, the maximal and minimal metabolic rates, metabolic scope, and the maximum swimming speed were measured. Results showed that the cost of maintenance of basal metabolic activities was particularly higher when n-3 HUFA food content was low. Moreover, fish exposed to high acclimation temperature combined with a low n-3 HUFA dietary level (LH20) exhibited a higher aerobic scope, as well as a greater expenditure of energy to reach the same maximum swimming speed as other groups. This suggested a reduction of the amount of energy available to perform other physiological functions. This study is the first to show that the impact of lowering n-3 HUFA food content is exacerbated for fish previously acclimated to a warmer environment. It raises the question of the consequences of longer and warmer summers that have already been recorded and are still expected in temperate areas, as well as the pertinence of the lowering n-3 HUFA availability in the food web expected with global change, as a factor affecting marine organisms and communities.