Induction of spermatogenesis in 1+ year-old greater amberjack (Seriola dumerili) after administration of single-chain recombinant gonadotropins

Introduction

Greater amberjack (Seriola dumerili) begin to reach sexual maturity after 4 years, and the age can vary in different geographical locations (Kožul, Skaramuca, Glamuzina, Glavić, & Tutman, 2001). A long period of time is required, as only at five to six years of age all the population is considered fully mature. Therefore, shortening the time between generations by advancing the time of puberty might bolster future production (Levavi- Sivan, Bogerd, Mañanós, Gómez, & Lareyre, 2010; Molés et al., 2020). We report, here, the in vivo efficacy of recombinant gonadotropin-based therapies in promoting gonadal development in 1+ year old, pre-pubertal greater amberjack.

Materials and Methods

An experiment was conducted to determine the minimum effective dose for the administration of recombinant Seriola dumerili single-chain gonadotropins (rsdGths), follicle-stimulating (rsdFsh), and luteinizing (rsdLh) hormones. Sixteen-month-old hatchery-produced F1 pre-pubertal male and female greater amberjack (October 2021) were treated with one of three doses of rsdFsh (Saline, 5, 10 or 15 μg kg-1) or two doses of rsdLh (5 or 10 μg kg-1) over six and two weeks, respectively. At the end of rsdFSH administration, gonads were dissected to estimate the gonado-somatic index (GSI) and for histological investigation. Blood samples were obtained from fish treated with rsdFsh as well as from those treated with rsdLh at regular interval to measure sex steroid hormones. In a second experiment, 1+ year old fish were injected with the best doses of rsdFSH and rsdLH established in the previous study, once a week over a period of twelve weeks, during the natural reproductive season for the species (March-June 2022). As above, blood samples were collected regularly to monitor variations in the plasma levels of sex steroids. A week after the last rsdGths administration, fish will be euthanized and gonads collected for the estimation of the gonado-somatic index (GSI) and for histological investigation.

Results

In the Control group, the testes were dominated by spermatogonia, and spermatocytes represented a small fraction of the gonadal content. The administration of 10 and 15 μg kg-1 of rsdFsh stimulated proliferation and differentiation of germ cells and a significantly increased GSI values (Fig. 1). Sperm was found within the testes, and histology confirmed the presence of all spermatogenic stages and spermatozoa in the seminiferous lobules. Additionally, male plasma levels of testosterone (T) and 11-keto T differed among groups and with a significant increase after administration of 10 and 15 μg kg-1 of rsdFsh.

Contrary to the males, no differences were found in the GSI values of females between groups. However, the mean diameter of the largest primary oocytes differed significantly, with a 27 and 29% increased size in females treated with 10 and 15 μg kg-1, respectively. Although ovarian lamellae filled with oogonia and primary oocytes were found in females from all groups, micrographs of ovarian sections displayed a higher content of oocytes in the perinuclear stage in the ovaries of females treated with 10 and 15 μg kg-1. Significantly increased plasma levels of T were also found in females given 10 and 15 μg kg-1 of rsdFsh, but no differences were detected in the plasma levels of 17β-Estradiol. Finally, no differences were found in plasma sex steroid levels in fish after injection with rsdLh.

Collection of experimental data from the second experiment in underway, and the results will be presented at the conference.

Discussion

The administration of rsdFsh only, was effective to induce gonadal development and early maturation in pre-pubertal male greater amberjack. Although oogenesis was not achieved in females, the increase in mean diameter of primary oocytes indicates that a longer administration of recombinant Fsh would have been beneficial for further development of the gonads. All together these results demonstrated the biological potency of rsdFsh, and that it can be used to induce gonadal development in immature greater amberjack, therefore shortening the onset of puberty. The response to a combined usage of rsdLh and rsdFsh in plasma steroid levels and gonadal development is currently under examination, and based on the results, the appropriate doses and timing for administration will serve the purpose of implementing protocols for full maturation on pre-pubertal fish during the regular reproductive season.