Challenges of sperm cryopreservation in transferring heat adaptation of corals across ocean basins.

Reef-building corals live very close to their upper thermal limits and their persistence is imperiled by a rapidly warming climate. Human interventions may be used to increase the thermal limits of sensitive corals by cross-breeding with heat-adapted populations. However, the scope of breeding interventions is constrained by regional variation in the annual reproductive cycle of corals. Here we use cryopreservation technology to overcome this barrier and cross-breed conspecific coral populations across ocean basins for the first time. During regional spawning events, sperm samples were cryopreserved from populations of the widespread Indo-Pacific coral, , from the southern Persian Gulf (maximum daily sea surface temperature of 36 °C), the Oman Sea (33 °C), and the central Great Barrier Reef (30 °C). These sperm samples were thawed during a later spawning event to test their ability to fertilize freshly spawned eggs of colonies from the central Great Barrier Reef. Average fertilization success for the Persian Gulf (9%) and Oman Sea (6%) sperm were 1.4-2.5 times lower than those for the native cryopreserved sperm from Great Barrier Reef (13-15%), potentially due to lower sperm quality of the Middle Eastern sperm and/or reproductive incompatibility between these distant populations. Overall, fertilization success with cryopreserved sperm was low compared with fresh sperm (>80%), likely due to the low motility of thawed sperm (≤5%, reduced from 50% to >90% in fresh sperm). To evaluate whether cross-bred offspring had enhanced thermal tolerance, the survival of larvae sired by Persian Gulf cryopreserved sperm, Great Barrier Reef cryopreserved sperm, and Great Barrier Reef fresh sperm was monitored for six days at ambient (27 °C) and elevated (33 °C) temperature. Against expectations of thermal tolerance enhancement, survival of larvae sired by Persian Gulf cryopreserved sperm was 2.6 times lower than larvae sired by Great Barrier Reef fresh sperm at 33 °C (27% versus 71%), but did not differ at 27 °C (77% versus 84%). This lack of enhanced thermal tolerance was unlikely due to outbreeding depression as survival was equally poor in larvae sired by Great Barrier Reef cryopreserved sperm. Rather, follow-up tests showed that cryoprotectant exposure during fertilization (0.1% DMSO) has a negative effect on the survival of larvae which is exacerbated at elevated temperature. Collectively, our findings highlight challenges of breeding corals for enhanced thermal tolerance using cryopreserved sperm, which may be overcome by methodological advances in the collection and preservation of high-quality motile sperm and minimizing the exposure time of eggs to cryoprotectants.