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12 pages, 6 figures, supporting information online at https://www.pnas.org/lookup/suppl/doi:10.1073/pnas.2112660118/-/DCSupplemental.-- Data Availability: RNA-Seq and .xls tables data have been deposited in a publicly accessible database: SEXTAN (https://sextant.ifremer.fr/record/5cb1485f-385e-46e9-9cc1-4b4d77802183/). All other study data are included in the article and/or supporting information, In most animals, sex determination occurs at conception, when sex chromosomes are segregated following Mendelian laws. However, in multiple reptiles and fishes, this genetic sex can be overridden by external factors after fertilization or birth. In some species, the genetic sex may also be governed by multiple genes, further limiting our understanding of sex determination in such species. We used the European sea bass (Dicentrarchus labrax) as a model and combined genomic (using a single nucleotide polymorphism chip) and transcriptomic (RNA-Sequencing) approaches to thoroughly depict this polygenic sex determination system and its interaction with temperature. We estimated genetic sex tendency (eGST), defined as the estimated genetic liability to become a given sex under a liability threshold model for sex determination, which accurately predicts the future phenotypic sex. We found evidence that energetic pathways, concerning the regulation of lipids and glucose, are involved in sex determination and could explain why females tend to exhibit higher energy levels and improved growth compared to males. Besides, early exposure to high-temperature up-regulated sox3, followed by sox9a in individuals with intermediate eGST, but not in individuals showing highly female-biased eGST, providing the most parsimonious explanation for temperature-induced masculinization. This gonadal state was maintained likely by DNA methylation and the up-regulation of several genes involved in histone modifications, including jmjd1c. Overall, we describe a sex determination system resulting from continuous genetic and environmental influences in an animal. Our results provide significant progress in our understanding of the mechanisms underlying temperature-induced masculinization in fish, The study was supported by the European Maritime and Fisheries Fund (3S, Seabass Sex and Stress, Grant No. 4320175237) allocated to B.G. Production of the fish benefited from AQUAEXCEL2020 Transnational access grant “Transsexbass” to F.P. and M.V. Research at the F.P. laboratory was supported by Spanish Ministry of Science Grant No. PID2019-108888RB-I00., With the institutional support of the ‘Severo Ochoa Centre of Excellence’ accreditation (CEX2019-000928-S)

In European sea bass (Dicentrarchus labrax), as in many other fish species, temperature is known to influence the sex of individuals, with more males produced at relatively high temperatures. It is however unclear to what extent growth or stress are involved in such a process, since temperature is known to influence both growth rate and cortisol production. Here, we designed an experiment aiming at reducing stress and affecting early growth rate. We exposed larvae and juveniles originating from both captive and wild parents to three different treatments: low stocking density, food supplemented with tryptophan and a control. Low stocking density and tryptophan treatment respectively increased and decreased early growth rate. Each treatment influenced the stress response depending on the developmental stage, although no clear pattern regarding the whole-body cortisol concentration was found. During sex differentiation, fish in the low-density treatment exhibited lower expression of gr1, gr2, mr, and crf in the hypothalamus when compared to the control group. Fish fed tryptophan displayed lower crf in the hypothalamus and higher level of serotonin in the telencephalon compared to controls. Overall, fish kept at low density produced significantly more females than both control and fish fed tryptophan. Parents that have been selected for growth for three generations also produced significantly more females than parents of wild origin. Our findings did not allow to detect a clear effect of stress at the group level and rather point out a key role of early sexually dimorphic growth rate in sex determination.

The European sea bass (Dicentrarchus labrax) exhibits female-biased sexual size dimorphism (SSD) early in development. New tagging techniques provide the opportunity to monitor individual sex-related growth during the post-larval and juvenile stages. We produced an experimental population through artificial fertilization and followed a rearing-temperature protocol (~16°C from hatching to 112 days post-hatching, dph; ~20°C from 117 to 358 dph) targeting a roughly balanced sex ratio. The fish were tagged with microchips between 61 and 96 dph in five tagging trials of 50 fish each; individual standard length (SL) was recorded through repeated biometric measurements performed between 83 to 110 dph via image analyses. Body weight (BW) was modelled using the traits measured on the digital pictures (i.e. area, perimeter and volume). At 117 dph, the fish were tagged with microtags and regularly measured for SL and BW until 335 dph. The experiment ended at 358 dph with the sexing of the fish. The sex-ratio at the end of the experiment was significantly in favor of the females (65.6% vs. 34.4%). The females were significantly longer and heavier than the males from 103 dph (~30 mm SL, ~0.44 g BW) to 165 dph, but the modeling of the growth curves suggests that differences in size already existed at 83 dph. A significant difference in the daily growth coefficient (DGC) was observed only between 96 and 103 dph, suggesting a physiological or biological change occurring during this period. The female-biased SSD pattern in European sea bass is thus strongly influenced by very early growth differences between sexes, as already shown in previous studies, and in any case long before gonadal sex differentiation has been started, and thus probably before sex has been determined. This leads to the hypothesis that early growth might be a cause rather than a consequence of sex differentiation in sea bass.

Temperature‐dependent sex determination (TSD) can be observed in multiple reptile and fish species. It is adaptive when varying environmental conditions advantage either males or females. A good knowledge of the thermosensitive period is key to understand how environmental changes may lead to changes in population sex ratio. Here, by manipulating temperature during development, we confirm that cold temperature (16°C) increases the proportion of fish that develop as females in European sea bass (Dicentrarchus labrax) until 56 days posthatching, but show that it has an opposite effect at later stages, with the proportion of males reaching ~90% after 230 days at 16°C. This is the first observation of opposite effects of temperature at different time periods on the sex ratio of a vertebrate. Our results highlight the potential complexity of environmental effects on sex determination., In the European sea bass, a marine fish species with a polygenic sex‐determination system, we show that exposure to low temperature has opposite effects on sex ratio. Early exposure favors female sex determination, while late exposure favors male sex determination.

International audience; Changes in ocean water temperature associated with global climate change are bound to enormously affect fish populations, with potential major economic consequences in the aquaculture and fisheries industries. A link between temperature fluctuations and changes in fish stress response is well established. In this study, we aimed to assess the effects of a short- (4 days) or a long-term (4 months) exposure to warm temperature in the stress physiology of European sea bass (Dicentrarchus labrax) larvae and juveniles. First, cortisol (i.e. the main stress hormone in fishes) analysis was used to confirm that a steady and short-term elevation of temperature acts as a physiological stressful event in these fish, and cortisol release is indeed above a metabolic increase linked to temperature. Moreover, our results verified that measurement of cortisol released into the water can be reliably employed as a non-invasive indicator of acute thermal stress in experimental conditions. Secondly, the different effects on the genetic cascade underlying the stress response between long-term low or high thermal treatments were evaluated at two larval development stages via candidate-gene and whole-transcriptome approaches. Interestingly, opposite expression for some key stress genes (nr3c1, nr3c2 and hsd11b2) were observed between developmental stages, highlighting the distinct adaptive mechanisms controlling the primary and secondary responses to a stressor. Surprising expression patterns for some understudied genes involved in the stress axis were also revealed, including crhr1, mc2r, mc5r, trh or trhr, which should be further explored. Finally, evaluation of cortisol content in scales was successfully used as a biomarker of chronic thermal stress, with 10x more cortisol in fish kept at 21 ◦C vs 16 ◦C after 4 months, supporting the gene expression results observed. The use of such a method as a proxy of long-term stress, unprecedented in the literature, holds a vast array of applications in further research, in particular, in the investigation of the impact of global warming on wild fish populations.

International audience; Individual tagging is key to a better understanding of early life stages in fish. Very small RFID transponder microchips (500 x 500 x 100 mu m, 82 mu g) are now available. The aim of this study was to develop a protocol to tag European sea bass (Dicentrarchus labrax L.) larvae from 61 days post-hatching (dph; standard length similar to 10 mm) to 96 dph (standard length similar to 28 mm) through intra-coelomic implantation of microchips. The suitability of such a tagging procedure was evaluated, with the purpose of determining the minimal fish age and body size for microchip tagging without adverse effects on survival and growth performance.We produced an experimental population composed by 50:50 normally pigmented larvae and albino larvae through artificial fertilization. Five tagging trials were performed over 35 days, in fish aged 61, 75, 83, 89 and 96 dph. Each time, 50 normally pigmented fish were tagged, while 50 albino fish were used as controls. Mortality was recorded daily, while biometric measurements were performed at 75, 83, 89, 96, 103 and 110 dph via image analysis.Microchip tagging was possible in larvae from an age of 75 dph (standard length similar to 20 mm), with satisfactory performance in terms of survival rate (between 84 and 98% 24 h after tagging) and growth rate, and without significant differences in comparison with the untagged controls. In contrast, tagging before 75 dph is not to be recommended, as the age group 61 dph was the most affected in terms of survival (only 62% of fish survived 24 h after tagging) and growth rate, showing significant differences compared to the untagged controls. The overall microchip reading success rate for the age groups throughout the experiment was 51.4%, the overall reading success rate at each biometric measurement was 48.2%, probably due to the change in orientation of the microchip inside the fish body cavity.The tagging protocol developed was then overall successful, albeit with a moderate reading success. Precocious tagging could allow the collection of new types of data (individual, longitudinal) related to larval development, behavioral studies, physiological and immunological investigations. Future tests could focus on the effects of tagging on baseline locomotion and behavior, as well as the suitability and the efficiency of intramuscular microchip tagging on larger fish.

Marie-Odile Blanc — Regional trajectories in Southern Africa in the context of global recession. This text aims at explaining the reactions of southern African economies to global recession and internal crisis. The volatility of primary product rates ; their sectoral effects and the strains on internal stability may compromise the prospects of regional growth. The influence of what the author terms here as the hegemonic South African pole and its relay-economies on the region, is analysed from the asymmetrical configuration of exchanges with southern Africa., Blanc Marie-Odile. Les trajectoires régionales en Afrique australe dans un contexte de récession mondiale. In: Tiers-Monde, tome 43, n°169, 2002. Les chemins de l'intégration régionale. pp. 71-90.