068 Challenges of creating a frozen repository for coral

Over the next 30 to 40 years, rising ocean temperatures and increased ocean acidification have the potential to overpower the vast majority of current in situ coral conservation efforts, resulting in severe loss of reef biodiversity. If this occurs, ex situ coral conservation incorporating cryopreservation may provide an important means for successful reef diversification. To build new tools for the continued protection and propagation of coral, an international group of coral and cryopreservation scientists known as the Reef Recovery Initiative joined forces. The outcome was the creation of the first frozen bank for Australian, Caribbean and Hawaiian coral, containing 8 important reef-building species including, Acropora tenuis, Acropora millepora, Acropora loripes, Platygyra daedalea, Platygyra lamolina, Fungia scutaria, Acropora palmata and Acropora cervicornis. Most species of coral are hermaphrodites and reproduce sexually by producing lipid-rich egg/sperm bundles. Generally, self-fertilization is low. A simple, uniform freezing method (10% Me2SO in seawater with a freezing rate of 20°C/min) was devised to cryopreserve the sperm of all these species. Coral reproduction is driven by access to sunlight, which powers the intracellular algae to produce complex sugars for the coral. In turn, the coral use these sugars to build lipids needed for the germplasm. Even though we found a unified method for sperm cryopreservation in coral there were physiological challenges. For example, sperm production across the complex web of polyps in single genetic individuals varied. Specifically, in 2011 egg/sperm bundles with more eggs produced more motile sperm. This variation may have been caused by polyp age, prior damage, access to sunlight or a combination of factors. An additional challenge was a ‘night-of-preservation’ effect whereby some nights the sperm preservation from pooled colonies produced widely differing post-thaw motility, as well as cryopreservation and fertilization success. Finally, when this simple sperm cryopreservation method was used on the Hawaiian acroporid, Montipora captitata, it consistently produced low post-thaw motility and no fertilization success. This species was found to have a toxin in the adult tissue that it transferred to the eggs and possibly the sperm prior to reproduction. Therefore, the release of this toxin from slightly damaged sperm cells may have impacted these post-thaw results, and may ultimately inhibit the successful cryopreservation of this species. Finally, coral larvae have not been successfully cryopreserved because of their exquisite chilling sensitivity. Nevertheless, pluripotent 8-cell embryos were successfully dissociated, frozen and thawed with 50 to 90% intact cells post-thaw in all species tested. Frozen and thawed coral sperm can now be used to create new coral, as they develop, settle and take up their intracellular algae. In the near future, we may have a blueprint to move our work from the laboratory to the reefs to develop collaborative, practical conservation management tools to secure reef biodiversity.Source of funding: This work was supported by the Roddenberry Foundation, Matthew Frank Foundation, Taronga Conservation Society Australia, Australian Institute of Marine Science, Anela Kolohe Foundation, Hawaii Institute of Marine Biology and the Smithsonian Institution.Conflict of interest: None [email protected]