Optimised donor plant nutrition improves cryopreservation of shoot-tips of an endangered species - Gossia fragrantissima (Myrtaceae)

Abstract Gossia fragrantissima (sweet myrtle) is an endangered Australian rainforest tree threatened by habitat loss and the invasive fungal disease myrtle rust. Due to limited wild populations and poor seed production, this study explored cryopreservation of shoot-tips as a long-term conservation strategy for preserving the remaining germplasm. We have previously shown that response surface modelling could deliver a significantly improved basal nutrient medium for G. fragrantissima shoot culture. Here for the first time, successful cryopreservation of G. fragrantissima apical shoot tips was achieved using an optimised droplet vitrification protocol for two genotypes. While one genotype (0124) showed high regeneration (> 90%) across all treatments, a second genotype (0119) was comparatively sensitive to cryopreservation. We showed that in this cryo-sensitive genotype, an RSM-optimised basal medium supported significantly enhanced cryopreservation outcomes compared to material cultured on the commercially available Woody Plant Medium (WPM; P < 0.05). On the RSM-optimised medium, genotype 0119 achieved up to 74% post-cryopreservation regrowth, and 0124 up to 97%. Comparatively, the highest regeneration rates achieved on WPM were 40% and 90%, respectively. This is the first study to show successful cryopreservation of any species from the Gossia genus, which contains other endangered Australian species as well as unique metal hyperaccumulator species. This study revealed that nutritional conditions in culture can be a decisive factor for cryopreservation outcomes and provides a good model for future in vitro conservation development for other threatened plants.