Introduction
Climate change is a serious concern for the aquatic environment, as it alters the physical and chemical properties of seawater causing negative impacts on fish. Among its many effects, the increase of atmospheric greenhouse gases has led to a rise in average sea surface temperatures. Fish are poikilothermic and are therefore extremely sensitive to changes in environmental temperature. E xposure of adult fish to warm temperatures is known to impair reproduction, although the long-term reproductive impact during early life stages remain poorly understood. The European sea bass (Dicentrarchus labrax L.) is a marine species of high commercial value, in which a significant proportion of individuals exhibit precoc ious puberty during the grow-out phase, when they are reared to marketable size . A situation that aquaculture producers want to prevent . The goal of this study was to evaluate the effect of high sea water rearing temperature on the onset of early puberty in th is species. G rowth and gonad maturation in juvenile 1-year-old male and 2- year-old female European sea bass were analysed, and its consequences in productive terms were considered.
Materials and methods
Juvenile sea bass (initial body weight (BW) of 14.0 ± 4.1 g) were provided by Piscialba-Piscifactorías Albadalejo S.L. (Murcia, Spain). Eight-month-old fish were sized and weighed, and the smallest 85% of the animals were considered potentially male fish (n = 510 fish) whereas the largest 15% were considered potentially female fish (n = 90). Potential male fish were distributed in six 500-l circular fiberglass tanks (n=85 fish/tank) whereas potential female fish were distributed in two 500-l circular fiberglass tanks (n = 45 fish/tank). After an acclimatization period of 30 days, two experimental groups for each sex were organized in triplicate in males and duplicate in females : control group (CG) that was maintained under natural seawater temperature conditions at the I ATS facilities (40°08′15″ N; 0°10′12 ″ E) and high- temperature group (HTG) that was maintained at a temperature that was 3-4 ºC above that of the CG . The survival rate as well as BW and fork length (FL) of fish was periodically measured. B lood samples were collected, and t he plasma was stored at -20ºC until hormonal analysis . In the case of males, in order to assess the testicular growth and the proportion of early maturing fish , 4-10 animals per group were sacrificed from July to February during their first year of life. T he ovarian growth and the incidence of early onset puberty was assessed in 4-10 f emales per group that were sacrificed from October to February during their second year of life. Gonad was removed and weighed to determine the gonadosomatic index (GSI) and s mall pieces of tissue were fixed for histological analysis and assessment of the stages of testicular and ovarian development . The percentage of early maturing males was also evaluated from November to onwards by applying gentle hand pressure to the abdomen of all fish. Circulating plasma levels of follicle stimulating-hormone (Fsh ) and its content in the pituitary were evaluated in both sexes. Plasma l evels of testosterone (T), 11-ketotestosterone (11-KT) and estradiol (E2 ) were also analysed in males, whereas levels of E2 and vitellogenin (Vtg) were analysed in females.
Results
After five months of experiment, male sea bass reared at a high temperature (HTG) showed a lower survival rate than that of the CG (66% and 96%, respectively). M ales in HTG was approximately 12.61 % lower in weight and 3.14 % smaller in length than that of CG at 17 months of age. The condition factor varied between 1. 01 and 1. 81 in fish at HTG, while it was around 1.02-2.14 in the CG. First significant increase of the GSI was observed after 4 months of experiment in fish of the CG (GSI= 0.47 ± 0.42% ) compared to males reared in HTG (GSI= 0.05 ± 0.03%). In February, during the tentative testicular growth period, GSI of the HTG was 1.66 ± 0.33% whereas values in the CG were significantly higher (2.74 ± 0.26%). The histological analysis of testis revealed clear differences between CG and HTG, where the percentages of gonadal stage III (m id recrudescence) in the CG reached 17% in October and 57% in November whereas the HTG group displayed 14.3% in December . In February, the percentages of gonadal stage IV (late recrudescence) in the CG was 89% and 11% in stage V (full spermiating testis) where all males in the HTG reached gonadal stage IV. Furthermore , the proportion of precociously spermiating males was higher in the CG than that of HTG from December to March, with a peak of 78% in the CG in comparison to 23% in the HTG in February. Of note , spermiation time lasted one month longer , until April, in the CG. Also, males in the HTG exhibited an increase of plasma and pituitary content of Fsh , whereas the levels of T and 11-KT remained unchanged in both experimental groups. In the case of females, no significant differences in growth performance were observed between CG and HTG. The condition factor ranged from 1. 19 and 1. 77 in the HTG and from 1.18-1. 61 in the CG group . Although values of GSI were higher in the CG compared to those of HTG, no significant differences were observed. In February, values of GSI of the HTG were 3.65 ± 1.04 % whereas those of the CG reached 6.58 ± 0.84 %. H istological analysis of ovaries revealed that the incidence of early onset puberty in females was similar in both groups. Accordingly, all females in the CG and HTG showed oocytes in late vitellogenesis in February, although the animals in the HTG group entered vitellogenesis one month later than those in the GC group. Finally, fe males in both experimental groups exhibited similar Fsh and E2 plasma levels, whereas the levels of vitellogenin increase later in the HTG group, aligning with the delayed vitellogenesis observed histologically.
Discussion
The se findings show that high sea water rearing temperatures reduce the long-term growth performance of juvenile male European sea bass during their first year of life. A lthough males can start spermatogenesis , fish in the HTG show a delay in the progression towards advanced testicular stages and a reduction in the time fish remain in spermiation. On the other hand, high sea water rearing temperatures d o not affect growth performance of female sea bass during their first two years of life and the incidence of early puberty is similar in both CG and HTG groups. Accordingly, high sea water rearing temperatures affect the reproductive axis of European sea bass at multiple levels, although t he occurrence of early sexual maturation persists and, in turn, its side effects need to be avoided in terms of production. F inally, high sea water rearing temperatures can impact spawning season, sometimes shortening it, thus influencing the life history traits of fish species and populations.
Acknowledgements
Research supported by THINKINAZUL/2021/042 project (MICIN, PRTR-C17. I1; Generalitat Valenciana) to A.G.-A.F. S.S. was supported by a research contract from Margarita Salas program from Universidad de Las Palmas de Gran Canaria.