Publications: Peer-reviewed journal articles (by staff)

Cryopreservation of sperm of the Pacific oyster (Crassostrea gigas): development of a practical method for commercial spat production

  • Adams SL,
  • Smith JF,
  • Roberts RD,
  • Janke AR,
  • Kaspar H,
  • Tervit HR,
  • Pugh PA,
  • Webb SC,
  • and King NG
1 January, 2004
CITATION

Adams SL, Smith JF, Roberts RD, Janke AR, Kaspar H, Tervit HR, Pugh PA, Webb SC, King NG 2001. Cryopreservation of sperm of the Pacific oyster (Crassostrea gigas): development of a practical method for commercial spat production. Aquaculture 242: 271-282.

ABSTRACT

This paper describes a simple method for cryopreserving sperm of the Pacific oyster (Crassostrea gigas Thunberg) in quantities suitable for commercial spat production. Experiments to refine. the cryoprotectant mixtures demonstrated the key role of trehalose. Trehalose alone (at 0.45 M final concentration) was an effective cryoprotectant. The addition of 2.5-15% dimethyl sulphoxide (DMSO) in combination with 0.45 M trehalose gave only modest improvement in fertility over trehalose alone (p=0.056). There was no significant difference in fertility among DMSO concentrations (p=0.611). Seawater (SW) without cryoprotectant gave very poor I results. but yielded some fertilization at very high sperm concentrations (7+/-1% at 10(7) sperm mL(-1), 21+/-2%at 3.2 x 10(7) sperm mL(-1), mean+/-S.E., n=3). The fertility of unfrozen sperm was 30- to 100-fold higher than that of sperm cryopreserved with DMSO and/or trehalose. For sperm cryopreserved in 4.5-mL cryovials, two simplified freezing methods gave fertilization rates equivalent to sperm cryopreserved by controlled rate freezing (p=0.386). These methods involved securing the cryovials to aluminium canes and then either placing them into a bath of methanol chilled with dry ice. or holding them on a floating rack 3 cm above liquid nitrogen. A third technique of plunging the cryovials directly into liquid nitrogen gave reduced and variable fertility relative to the methanol/dry ice bath method (p=0.032). The commercial applicability of the protocols was demonstrated on a batch of 30 million eggs. Fertilization with cryopreserved sperm yielded 81% fertilization, and larval rearing by normal commercial practises yielded 3.7 million settled spat, which was comparable to the 2.5 million spat from a parallel batch fertilized with unfrozen sperm. (C) 2004 Elsevier B.V All rights reserved.