title: Cryopreservation of Greenshell™ mussel (Perna canaliculus) trochophore larvae.
abstract: The Greenshell™ mussel (Perna canaliculus) is the main shellfish species farmed in New Zealand. The aim of this study was to evaluate the effects of cryoprotectant concentration, loading and unloading strategy as well as freezing and thawing method in order to develop a protocol for cryopreservation of trochophore larvae (16-20 h old). Toxicity tests showed that levels of 10-15% ethylene glycol (EG) were not toxic to larvae and could be loaded and unloaded in a single step. Through cryopreservation experiments, we designed a cryopreservation protocol that enabled 40-60% of trochophores to develop to D-larvae when normalized to controls. The protocol involved: holding at 0 °C for 5 min, then cooling at 1 °C min⁻¹ to -10 °C, holding for a further 5 min, then cooling at 0.5 °C min⁻¹ to -35 °C followed by a 5 min hold and then plunging into liquid nitrogen. A final larval rearing experiment of 18 days was conducted to assess the ability of these frozen larvae to develop further. Results showed that only 2.8% of the frozen trochophores were able to develop to competent pediveligers.
authors.affiliation: Departamento de Ecoloxía e Bioloxía Animal, Universidade de Vigo, Estrada Colwxio Universitario s/n, 36310 Vigo, Galicia, Spain. eparedes@uvigo.es
authors.forename: E
authors.initials: E
authors.lastname: Paredes
authors.forename: S L
authors.initials: SL
authors.lastname: Adams
authors.forename: H R
authors.initials: HR
authors.lastname: Tervit
authors.forename: J F
authors.initials: JF
authors.lastname: Smith
authors.forename: L T
authors.initials: LT
authors.lastname: McGowan
authors.forename: S L
authors.initials: SL
authors.lastname: Gale
authors.forename: J R
authors.initials: JR
authors.lastname: Morrish
authors.forename: E
authors.initials: E
authors.lastname: Watts
mesh_terms: D000818:Animals; D015925:Cryopreservation / Q000379:methods*; D003451:Cryoprotective Agents / Q000378:metabolism / Q000633:toxicity; D019855:Ethylene Glycol / Q000378:metabolism / Q000633:toxicity; D007814:Larva / Q000187:drug effects / Q000254:growth & development; D049883:Perna / Q000187:drug effects / Q000254:growth & development*; D014199:Trehalose / Q000378:metabolism / Q000633:toxicity
chemical_list: D003451:Cryoprotective Agents; D014199:Trehalose; D019855:Ethylene Glycol