The hypothesis assume the existence differences in bulls spermatozoa viability after thawing, characterized by higher values of motility and higher proportion of live spermatozoa, in case of prolonged equilibration and lower temperature freezing gradient. The aim of this thesis was to find the influence of different equilibration and freezing curve conditions to on spermatozoa viability after thawing. The theoretical part of the thesis summarizes available information about the bulls semen parameters, methods for its evaluation and especially the processing and long-term preservation.
Within the case study part of the thesis, during the years 2013-2015 the semen of the pre-selected group of 5 breeding bulls was being collected at the Sires insemination center. The group was uniform as to age, breed and housing and treatment conditions. After initial assessment the collected semen was divided into parts, which were then processed in different wals. A part of straws was processed by standard protocol, the other part was processed using different length of equilibration and freezing curve.
First part of the straws was equilibrated in a standard protocol for 120 minutes; subsequently half of the equilibrated straws was standardly frozen by the Direct Freezing method based on 3-phase freezing curve while the other half was frozen using the different 2-phase freezing curve. Second part of the straws was initially equilibrated for 240 minutes with the subsequent freezing, similarly as in the previous case, used 3-phase and 2-phase freezing curve. Frozen straws were then stored in liquid nitrogen container at -196 °C.
Spermatozoa motility was evaluated using CASA. The semen was further evaluated on the proportion of live and dead spermatozoa, using coloration of Eosin and Nigrosine, and on the sensitivity of the spermatozoa membranes, using HOS test, which detects the reaction of spermatozoa tail membrane to hypoosmotic conditions.
The obtained results indicated higher progressive spermatozoa motility (+5.57 %) and higher percentage of live spermatozoa (+4.47 %) following the application of the prolonged equilibration (240 min). When applying the length of equilibration 240 minutes, the values of total spermatozoa motility and HOS test were higher as well, though the differences were not statistically significant. When evaluate the effect of using different freezing curves on the parameters of the spermatozoa after thawing, higher values (average + 1.6 %) at almost all of these parameters using 3-phase freezing curve were found, the diferrences were not statistically significant.
Statistically significant differences among the bulls (P < 0.01) were found in total as well as progressive spermatozoa motility, in the proportion of living spermatozoa and in value of the HOS test - the individuality among the bulls is therefore evident. While evaluating the characteristics of spermatozoa movement by CASA, statistically significant differences among the bulls were confirmed at VCL, VSL, VAP, LIN, STR, WOB (P < 0.01), also between the diferrent length of equilibration (P < 0.01), but were not statistically significant when using diferrent freezing curves.
| Identifer | oai:union.ndltd.org:nusl.cz/oai:invenio.nusl.cz:259931 |
| Date | January 2016 |
| Creators | Beránková, Monika |
| Contributors | Stádník, Luděk, Zuzana, Zuzana |
| Publisher | Česká zemědělská univerzita v Praze |
| Source Sets | Czech ETDs |
| Language | Czech |
| Detected Language | English |
| Type | info:eu-repo/semantics/masterThesis |
| Rights | info:eu-repo/semantics/restrictedAccess |
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