37 Survival rate of immature oocytes postcryopreservation by solid surface vitrification, conventional straw vitrification, or conventional slow-freezing method
M. L. Mphaphathi A B , S. M. Sithole A , M. D. Sebopela A , M. R. Ledwaba A , M. A. Thema A , T. L. Magopa A , H. A. O’Neill B and T. L. Nedambale AA Agricultural Research Council, Animal Production, Germplasm Conservation and Reproductive Biotechnologies, Pretoria, RSA
B University of the Free State, Department of Animal, Wildlife and Grassland Sciences, Bloemfontein, RSA
Reproduction, Fertility and Development 35(2) 144-145 https://doi.org/10.1071/RDv35n2Ab37
Published: 5 December 2022
© 2023 The Author(s) (or their employer(s)). Published by CSIRO Publishing on behalf of the IETS
Successful oocyte cryopreservation is important for establishing animal germplasm banks and animal fertility and biodiversity. The objective of the study was to compare survival rate of cattle immature oocytes following cryopreservation by solid surface vitrification (SSV), conventional straw vitrification (CSV), and conventional slow freezing (CSF). Heterogeneous cattle ovaries were collected from abattoir. The different cryoprotectant (CPA) concentrations (0, 5, 10, 15% followed by 10, 20, and 30%, stepwise increased CPAs) of dimethyl sulfoxide (DMSO) + ethylene glycol (EG) were prepared in M199. During CSF of oocytes (n = 232), a programmable freezer was used. For the SSV, the device was placed inside a Styrofoam box with liquid nitrogen (LN2) before vitrification of the oocytes (n = 233). During CSV method for oocytes (n = 238), mini straws were loaded with oocytes. The straws were pre-cooled by placing them horizontally on a Styrofoam rack exposed above the LN2 vapour. Cryopreserved oocytes were then subjected into warming or thawing media and then incubated in maturation medium for 22 h. The oocytes’ morphology and polar body extrusion was evaluated. These observations were repeated 12 times per treatment at different CPA concentrations. Treatment means were compared using a least significant difference test. The oocytes with zona cracked (9.0 to 46.1%) were noticed in control groups (cryopreserved without CPA) only (P < 0.05). Splitting of oocytes were on SSV EG + DMSO groups (ranging from 9.8 to 29.0%) and control CSV (7.5%; P > 0.05). Changes in oocytes’ shape were recorded in all the cryopreservation methods and different level of CPA % (P < 0.05). The changes in oocytes’ shape (control, no CPA) for CSF (35.8%), SSV (42.1%) and CSV (82.6%) methods were recorded (P < 0.05). Furthermore, the changes in oocytes’ shape (15% EG + 15% DMSO) for CSF (36.5%), SSV (10.1) and CSV (5.3) methods were recorded in different CPA concentrations (P < 0.05). In addition, the leakage of oocytes’ contents ranged from 0.0 to 28.9% (P < 0.05). The highest percentage of oocytes with polar body extrusion was 22.5% and was recorded from the CSV method (15% DMSO + 15% EG) and was not observed on the CSF and SSV methods (P < 0.05). In conclusion, the CSV method was the best method to cryopreserve the cattle immature oocytes and, subsequently, polar body extrusion.
The authors acknowledge the Agricultural Research Council and National Research Foundation for funding.
