Finding an effective freezing protocol for Turkey semen: Benefits of ficoll as non-permeant cryoprotectant and 1:4 as dilution rate

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

Validation of the Turkey semen cryopreservation by evaluating the effect of two diluents and the inseminating doses

This study was designed to test the fertilizing ability of cryopreserved turkey semen, and here, two experiments were performed: an in vitro analysis to assess the effects of Tselutin and Lake diluents and an in vivo test to determine the fertility and hatching rates by also studying the feat of three insemination doses (250, 400 and 600 7 106 sperm/hen). Pooled semen samples were diluted with Tselutin or Lake extender which contained 20% of dimethylsulfoxide and 1 mM of Ficoll at final sperm concentration of 3 7 109 sperm/mL. Thereafter, semen was packaged into straws and frozen on liquid nitrogen. The post-thaw sperm quality was evaluated considering motility (computer-aided sperm analysis\u2014CASA system) and membrane integrity (flow cytometry). Significantly higher values of progressive motility and some kinetic parameters in semen frozen with Lake were found. When we compared the extenders in vivo, no significant effects were detected, whilst sperm concentration significantly affected both fertility and hatching rates, with the best results obtained with the sperm concentration of 400 7 106 sperm/hen. From the results obtained, it emerged that the extender type only affected sperm motility characteristics, not the fertilizing ability of frozen-thawed semen, while inseminating dose markedly affected fertility and hatching rates

Italian semen cryobank of autochthonous chicken and turkey breeds: a tool for preserving genetic biodiversity

The creation of genetic resource cryobanks provides a crucial link between in situ and ex situ techniques to improve the efficiency of conservation programs. Aim of the present review is to describe all the activities developed for the implementation of the first Italian Semen Cryobank of Autochthonous Chicken and Turkey Breeds. These activities can be classified into three main topics: (1) identification of species-specific semen freezing/thawing reference procedures; (2) drafting Standard Operative Procedures (SOP) for the implementation of the semen cryobank; (3) storage of semen doses from Italian chicken and turkey breeds to establish the cryobank. Several trials have been developed to identify a specie-specific semen cryopreservation protocol for chickens and turkeys. The major results are reviewed and a final reference protocol described. Taking into consideration the FAO guidelines for cryoconservation of animal genetic resources, SOP were drafted with the aim to provide technical guidance and logistical support on the choice of priority breeds, selection of birds for semen production, infrastructures and storage sites, birds and semen management, cryopreservation process and doses traceability. Lastly, the Italian Semen Cryobank was created. A total of 112 semen doses from 22 cockerels of three breeds, and 74 doses from 12 turkey males of three breeds were stored in the Cryobank. Breed specific semen quality parameters assessed before and after cryopreservation are reported. The described activities provide information and tools useful for the implementation of semen cryobanking in avian species and might be transferred also to other species after appropriate adaptations.HIGHLIGHTS Implementation of the first Italian Semen Cryobank of Autochthonous Chicken and Turkey Breeds Drafting Standard Operative Procedures provides technical guidance and logistical support on the design and establishment of the cryobank Semen cryobank is a precious genetic reservoir and could be useful to safeguard genetic variability in small population in vivo conserved