Artificial insemination (AI) with sex-sorted bottlenose dolphin spermatozoa
provides female calves for obtaining more cohesive social groups and optimum genetic
management of captive populations. However, distance of animals to the sorting facility
represents a limit to the procedure. Although one bottlenose dolphin calf has been born
using spermatozoa from frozen-thawed, sorted and recryopreserved spermatozoa,
critical evaluation of the steps involved in this process is required to maximize its
efficiency for future AIs and expansion of the technology to other species.
Two experiments were designed to determine the efficiency of the sorting
process and the quality of frozen-thawed bottlenose dolphin spermatozoa during sorting
and recryopreservation. In experiment 1, the effect of two washing media (with and
without 4 percent egg yolk, v/v) following density gradient centrifugation (DGC) on sperm
recovery rate and in vitro characteristics of cryopreserved spermatozoa was examined.
In experiment 2, cryopreserved semen was used to compare the effects of two
recryopreservation methods (conventional straw freezing and directional freezing) on in
vitro sperm characteristics of control (non-sorted) and sorted spermatozoa. Egg yolk
supplementation of the washing medium in experiment 1 did not influence (P > 0.05) the sperm recovery rate, however, sperm motility parameters and viability were improved (P
< 0.05). For Experiment 2, motility parameters and viability were influenced by stage of
sex-sorting process, sperm type (non-sorted and sorted) and freezing method (P <
0.05). After recryopreservation, sorted spermatozoa frozen with the directional freezing
method maintained higher (P < 0.05) motility parameters over the 24 h incubation
period compared to spermatozoa frozen using straws. Quality of sperm DNA of nonsorted
spermatozoa, as assessed by the SCSA, remained unchanged throughout the
process. However, a possible interaction between Hoechst 33342 and acridine orange
was observed in sorted samples. After recryopreservation, viability of sorted
spermatozoa was higher (P < 0.05) than that of non-sorted spermatozoa across all time
points. The percentages of viable spermatozoa determined by light (eosin-nigrosin) and
fluorescence microscopy (propidium iodide) techniques were correlated (R^2=0.79, P <
0.001).
Collective results indicate that bottlenose dolphin spermatozoa undergoing
cryopreservation, sorting and recryopreservation are of adequate quality for use in AI.
| Identifer | oai:union.ndltd.org:tamu.edu/oai:repository.tamu.edu:1969.1/148447 |
| Date | 14 March 2013 |
| Creators | Montano Pedroso, Gisele 1981- |
| Contributors | O'Brien, Justine K, Kraemer, Duane C |
| Source Sets | Texas A and M University |
| Detected Language | English |
| Type | Thesis, text |
| Format | application/pdf |
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