First instance of settlement by cryopreserved coral larvae in symbiotic association with dinoflagellates

Abstract

Coral reefs are suffering on a global scale due to human impacts, thereby necessitating cryopreservation efforts. The objective of this study was to develop a suitable vitrification and laser warming protocol for larvae of the scleractinian coral Seriatopora caliendrum, which inherit their dinoflagellate algal symbionts vertically. Toxicity experiments were conducted with the cryoprotectants (CPAs) ethylene glycol (EG), propylene glycol (PG), dimethyl sulfoxide (DMSO), glycerol (GLY), and methanol (METH; listed in order from least to most toxic), and larvae were subjected to vitrification and laser warming using 2 M EG + 1 M PG and 2 M EG + 1 M DMSO. Vitrification and laser warming (300 V, 10 ms pulse width, 2 mm beam diameter) using a vitrification solution of 2 M EG + 1 M PG, 40% w/v Ficoll, and 10% v/v gold nanobars (GNB) at a final concentration of 1.2 × 1018 GNB/mL and a characteristic wavelength of 535 nm resulted in larvae with vitality and settlement percentages of 55 and 9%, respectively. This represents the first successful instance of cryopreservation of coral larvae that proceeded to settle upon warming, and suggests that the vitrification and ultra-fast laser warming approach may be applicable to other threatened marine species.

Cryopreservation and Cryobanking of Cells from 100 Coral Species

“…However, oocyte cryopreservation with MeOH requires vitrification with EG and propylene glycol [ 58 , 59 ] because high concentrations of MeOH can be toxic to oocytes. Nevertheless, attempts to cryopreserve asymbiotic [ 60 ] and symbiotic [ 61 , 62 ] coral larvae through vitrification and laser nanowarming have been successful.…”

“…Feuillassier et al (2014) [ 65 ] also cryopreserved coral ( P. damicornis ) apices using 0.2 M sucrose + 0.75 M DMSO + 0.75 M MeOH + 0.75 M EG. The larvae of F. scutaria and S. caliendrum have been preserved with 10% PG + 5% DMSO + 1 M trehalose and 2 M EG + 1 M propylene glycol [ 60 , 61 ]. Finally, coral Symbiodiniaceae (e.g., Symbiodinium , Breviolum , Cladocopium , Durusdinium , Fugacium , and Gerakladium species) have been cryopreserved with 2 M EG, 2 M MeOH, 1 M Gly, 1 M MeOH, and 1 M MeOH + 0.4 M sucrose [ 11 , 14 ].…”

“…However, EG was less effective in protecting P. damicornis tissue balls against CPA toxicity than MeOH, glycerol, and DMSO were [ 65 ]. EG combined with other CPAs has been used in vitrification solutions for J. juncea oocytes (Tsai et al, 2015) and symbiotic coral ( S. caliendrum and P. verrucosa ) larvae cryopreserved through vitrification and laser nanowarming [ 33 , 61 ].…”

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

“…Cryoprotectants are essential to eliminate ice formation when cooling cells to cryogenic temperatures but have concentration, exposure, tissue, and species-specific toxic effects which present ongoing challenges to cryopreservation success ( Best, 2015 ; Cirino et al, 2019 ). In corals, DMSO is the most widely used cryoprotectant and has generally been shown to have lower toxicity to coral sperm than alternatives ( e.g.…”

Cryopreservation of six Symbiodiniaceae genera and assessment of fatty acid profiles in response to increased salinity treatments

“…Successful cryopreservation of symbiotic dinoflagellates will provide a resource that can be used in research and the conservation of coral reefs 16 . Use of different cryopreservation techniques and appropriate amount of cryoprotectants have led to high cell survival of the Breviolum sp.…”