105. TRANSPLANTED GERM CELLS CAN COLONIZE THE GONADS OF SEXUALLY COMPETENT FISH AND PRODUCE FUNCTIONAL GAMETES

Abstract

Germ cell (GC) transplantation (GCT) has great potential for seed production and conservation of valuable germlines. Currently available approaches to GCT in fish rely on sophisticated equipment and skills for cell transplantation into the blastodisc of embryos and/or the peritoneal cavity of small, sometimes few millimeters long larvae. Moreover, transplanted individuals may take years to grow to maturity, adding to the cost of producing surrogate gametes. In this context, the use of intragonadal GCT into sexually competent recipients that have been experimentally depleted of endogenous GCs might overcome these constraints. Here we demonstrate the feasibility of xenogeneic GCT in sexually competent fish. Spermatogonial cells isolated from pejerrey (Odontesthes bonariensis, Atherinopsidae) donors were implanted surgically into the testes of congeneric Patagonian pejerrey (O. hatcheri) that were severely depleted of endogenous GCs by treatment with Busulfan (40 mg/kg) and elevated water temperature (25°C) (Fig. 1). Donor cell behavior inside the recipient gonads was tracked using fluorescent cell linkers (CFDA-SE and PKH-26) and showed that transplanted spermatogonial cells were able to migrate towards, settle and multiply at the blind ends (cortical region) of the seminiferous lobules. The presence of donor-derived sperm was confirmed by PCR in 20% of the surrogate Patagonian pejerrey fathers at 6 months and fertilization of pejerrey eggs with surrogate sperm produced 1.2-13.3% pure pejerrey offspring (Fig. 2). These findings indicate that transplantation of spermatogonial cells into sexually competent fish can shorten considerably the production time of surrogate gametes and offspring. Ongoing studies are examining (low-tech) refinements in the proposed approach, such as non-surgical transplantation of GCs through genital papilla, and the suitability of GCT for generation of female gametes, for which cryopreservation techniques have not yet been developed. The results obtained so far have been encouraging and these developments will make GCT invaluable for the timely rescue of fish species facing imminent extinction.