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. 2021 Jul 29;11:15428. doi: 10.1038/s41598-021-94197-2

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© The Author(s) 2021

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

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Post-thaw fitness parameters on larvae of Ruditapes philippinarum cryopreserved at trochophore, 48 and 72 h-old D-larval stages: (A) survival of cryopreserved trochophores; (B) shell size of developed D-larvae from cryopreserved trochophores; (C) survival of cryopreserved 48 h-old D-larvae; (D) shell size of developed D-larvae from cryopreserved 48 h-old D-larvae; (E) survival of cryopreserved 72 h-old D-larvae; (D) shell size of developed D-larvae from cryopreserved 72 h-old D-larvae. Larvae were cryopreserved using two CPA solutions: 10% ethylene–glycol (EG) + 0.4 M trehalose (TRE) and 10% prophylene-glycol (PG) + 0.4 M TRE (final concentrations), both prepared in filtered sea water (FSW). Increasing equilibrium times were tested on D-larvae. Results are expressed as mean normal D-larval percentage ± standard deviation, SD (n = 100, 3 replicates per treatment) or as box plots for shell size assessments (µm) considering mean value (mean ± standard deviation, SD) (n = 35, 3 replicates per treatment). Different letters show statistical differences with p˂0.05.