The Effects of Larval Cryopreservation on the Epigenetics of the Pacific Oyster Crassostrea gigas

Abstract

High mortalities and highly variable results during the subsequent development of post-thaw larvae have been widely considered as key issues restricting the application of cryopreservation techniques to support genetic improvement programs and hatchery production in farmed marine bivalve species. To date, few studies have been undertaken to investigate the effects of cryodamage at the molecular level in bivalves. This study is the first to evaluate the effect of larval cryopreservation on the epigenetics of the resultant progenies of the Pacific oyster Crassostrea gigas. The results show that the level of DNA methylation was significantly (p < 0.05) higher and lower than that of the control when the trochophore larvae were revived and when they developed to D-stage larvae (day 1 post-fertilization), respectively, but the level returned to the control level from day 8 post-fertilization onwards. The expression of the epigenetic regulator genes DNMT3b, MeCP2, JmjCA, KDM2 and OSA changed significantly (p < 0.05) when the trochophore larvae were thawed, and then they reverted to the control levels at the D- and later larval developmental stages. However, the expression of other epigenetic regulator genes, namely, MBD2, DNMT1, CXXC1 and JmjD6, did not change at any post-thaw larval developmental stage. For the newly thawed trochophore larvae, the amount of methylated H3K4Me1 and H3K27Me1 significantly changed, and the expression of all Jumonji orthologs, except that of Jumonji5, significantly (p < 0.05) decreased. These epigenetic results agree with the data collected on larval performances (e.g., survival rate), suggesting that the effect period of the published cryopreservation technique on post-thaw larvae is short in C. gigas.