Finding an Effective Freezing Protocol for Turkey Semen: Benefits of Ficoll as Non-Permeant Cryoprotectant and 1:4 as Dilution Rate

Simple Summary The most adopted biotechnology for the conservation of genetic resources in avian species is semen cryopreservation. Therefore, the identification of a reference cryopreservation procedure represents a key point for ensuring the long-term conservation of genetic diversity in birds, through the implementation of a semen cryobank. In this study, our goal was to discover an effective freezing protocol for Meleagris gallopavo in order to realize the first Italian semen cryobank of autochthonous chicken and turkey breeds within our project (TuBAvI). For this purpose, we investigated the effects of three non-permeant cryoprotectants (sucrose, trehalose, and Ficoll) and two dilution rates (1:2 and 1:4) on the in vitro cryosurvivability of turkey spermatozoa. After thawing, the best semen quality was found in semen frozen in the presence of Ficoll and diluted at a final rate of 1:4. This paper provides encouraging results, however further studies are programmed to standardize the semen cryopreservation protocol.Abstract The present study aimed to find an effective cryopreservation protocol for turkey semen through the combined use of dimethylsulfoxide (DMSO) and three non-permeant cryoprotectants (NP-CPAs), sucrose, trehalose, and Ficoll 70. In addition, the action of two dilution rates (1:2 and 1:4) were also investigated. Semen was processed according to two final dilution rates and the following treatments: Tselutin extender (TE)/DMSO (control), TE/DMSO + sucrose or trehalose 50, 100, 200, or 400 mM, and TE/DMSO + Ficoll 0.5, 0.75, 1, or 1.5 mM. In total 26 different combinations treatments were achieved. The diluted semen was filled up into straws and frozen on liquid nitrogen vapor. The post-thawing sperm quality was assessed by analyzing motility, membrane integrity, osmotic resistance, and DNA integrity. The results obtained revealed a significant effect of NP-CPA concentration on total and progressive motility, on most of the kinetic parameters, on membrane integrity and DNA integrity, while the post-thaw quality was less affected by dilution rate. The highest post-thaw quality for all sperm quality parameters assessed except curvilinear velocity (VCL) and DNA integrity were found in semen frozen with 1 mM Ficoll/1:4 (p < 0.05). Our findings provide an important contribution for the identification of a reference procedure for turkey semen cryopreservation, in order to create the first national avian semen cryobank.