Production of inbred larvae through self-fertilization using oocytes and cryopreserved sperm from the same individuals after sex reversal in eastern oyster Crassostrea virginica

The eastern oyster Crassostrea virginica can change sex which makes self-fertilization possible if sperm can be cryopreserved. In this study, small (similar to 1year old) and large (similar to 2-3years old) oysters were biopsied for sperm collection. Survival of the biopsied oysters after 1year was 50% for small oysters and 17% for large oysters. Oocytes were collected from sex-reversed females, and self-fertilized with cryopreserved sperm. Of the 24 cryopreserved samples, 14 individuals had 1% fertility when crossed with oocytes from unrelated females, indicating that the cryopreserved sperm had reduced fertility. The other 10 individuals had a fertility of 39 +/- 25% when crossed with oocytes from unrelated females (non-selfing), but showed a significantly lower success of self-fertilization (12 +/- 16%) (P=0.008), while aliquots of the same oocytes had a fertilization of 83 +/- 11% when crossing with fresh sperm. Larvae were produced at day 3 in the self-fertilized families (12-94% of the fertilized oocytes), and survived to eyed-larvae stage at days 11-14. Genotyping with 9 microsatellite markers confirmed that the larvae resulted from self-fertilization in four families. This study demonstrated the feasibility of creating self-fertilized inbred lines of oysters by use of non-lethal sperm collection and cryopreservation.