Sex differentiation and early sex identification of the greater amberjack Seriola dumerili reared in sea cages
Creators: name: Papadaki, Maria
name: Mandalakis, Manolis
name: Anastasiou, T.I.
name: Pouli, M.
name: Asderis, M.
name: Katharios, Pantelis
name: Papandroulakis, N.
name: Mylonas, Constantinos
Publisher: name: Zenodo
Year: 2023

description: Introduction
The greater amberjack Seriola dumerili is a fast-growing fish, for which commercial production has been recently achieved in the Mediterranean. The study of its sex differentiation process under captivity is essential, in order to investigate for possible sex-related growth differences and sex ratio fluctuations during on-growing in sea cages. Moreover, sex identification at an early age is very useful, in order to ensure optimal sex ratios and select specific female and male breeders for reproduction.
Materials and Methods
Fish used in the present study were produced from eggs obtained on June 26, in Argosaronikos Fish Farm S.A. (Salamina Island, Greece), after spawning induction of wild-caught breeders with gonadotropin releasing hormone agonist (GnRHa) implants (Fakriadis et al., 2020). Eggs were transferred to the facilities of the Institute of Marine Biology, Biotechnology and Aquaculture (Hellenic Center for Marine Research, HCMR) and reared until 50 days post-hatching (dph). Then, fish were moved to the pilot sea cages of HCMR at Souda Bay, Chania, Crete, Greece, where they were maintained until the end of the experiment.
Seven samplings were conducted from 101 until 408 dph (n=17-23), during which total length (TL) and wet weight (WW) were measured, and gonads and blood were collected, the former for histological analysis and the latter for the measurement of sex steroid (adrenosterone (Ad), androstenedione (Δ4), 11-ketotestosterone (11-ΚΤ), testosterone (Τ), estradiol (Ε2), progesterone (P4) and 17,20β-dihydroxyprogesterone (DHP)) and corticosteroid (cortisol and cortisone) plasma levels with the use of liquid chromatography/mass spectrometry (LC-MS/MS).
Results and Discussion
Growth of greater amberjack was similar for both sexes and females had the same size as males at all samplings. This finding suggests that both sexes should be equally preferable in aquaculture production. Moreover, the sex ratio in all the samplings was 1:1, meaning that no fluctuations are to be expected in the sex ratio of cage-reared populations and that there is no effect of the rearing method on the sex differentiation of the species.
As regards the histological description of sex differentiation, in females at 101 dph (14.98 ± 6.20 cm TL), the ovarian cavity was already formed, and germ cells were visible around it. At 198 dph (25.8 ± 0.14 cm TL) the first primary oocytes were visible. The typical ovarian structure with ovarian lamellae and occasional presence of primary oocytes was apparent at 260 dph (27.75 ± 1.89 cm TL), whereas complete ovary differentiation was achieved at 408 dph (41.22 ± 3.83 cm TL).
In males, at 101 dph the gonads contained mostly somatic cells and connective tissue and no germ cells were present (14.47 ± 6.60 cm TL). The first germ cells were apparent at 150 dph, when spermatocytes could be found in the gonads (24.71 ± 3.09 cm TL). The number of proliferating and differentiating germ cells increased at 198 and 260 dph (28

± 2.39 cm and 29.75 ± 2.82 cm TL, respectively) and the typical testicular structure featuring all types of male germ cells, was observed at 260 dph (28.63 ± 2.85 cm TL). This structure was obviously maintained until the last sampling (357 and 408 dph, 34.25 ± 2.98 cm and 41.7 ± 1.99 cm TL, respectively).
Of the nine measured hormones in greater amberjack females and males during the sex differentiation process, the ones that exhibited statistically significant changes in time were cortisol, Ad, Δ4, 11-KT, T and P4 in females and Ad, 11-KT, T, P4 and DHP in males. Cortisol and Ad exhibited lower values as the sex differentiation period was progressing in females; on the other hand, Ad, 11-KT, T and P4 in both sexes and DHP in males presented higher values at the end of the sex differentiation period.
The 11-KT concentration was significantly different between the two sexes at all samplings, except at 198 dph. The 11-KT/E2 ratio was significantly different between the sexes from the second sampling at 150 dph until the last sampling at 408 dph.
Conclusions
Hatchery-produced greater amberjack shows no sexual size dimorphism and the sex ratio in the cultured population was always 1:1, underlining that the early life rearing method did not have any influence on the process of sex differentiation. Sex identification could be conducted in juvenile fish from 150 dph, with the use of the 11-KT/E2 ratio. Moreover, using LCMS/MS, with just 200 μL of plasma, a large number of steroid hormones of the cholesterol metabolism pathway could be detected, enabling the study of the biochemical pathway involved in the teleost sex differentiation process. More studies are needed in order to decipher the exact role of each separate hormone in sex differentiation, both of greater amberjack and of other species.