Semen cryopreservation, laparoscopic artificial insemination and embryo transfer in goat in Thailand

Abstract

The study is aimed to do the research on semen cryopreservation, laparoscopic artificial insemination and embryo transfer in goats and provide valuable practical opportunities to improve reproductive efficiency and to enhance the genetic improvement. It composed of six experiments. Experiment I, II and III aimed at evaluation the effect of Equex STM Paste and glycerol supplemented in freezing extender on freezability and fertilizing ability. The semen qualities including motility, viability, morphology, acrosome integrity, membrane functional integrity and DNA integrity, were evaluated before processing and after cryopreservation. Computer-assisted-semen analysis (CASA) was used to characterize the sperm motion patterns during the thermal resistance test at 0, 1, 2 and 3 h post-thawing. The freezing medium containing 10% glycerol and 0.5% Equex STM Paste sufficiently protected goat spermatozoa against cryoinjury during freezing and thawing, and provided an acceptable pregnancy rate. Experiment IV, aimed to study the seasonal effect on semeinal characteristics and freezability of crossbred Saanen bucks. Overall, the fresh semen collected in summer and winter provided a higher in quality. The post-thaw semen qualities were not significant different except for motility characteristics and plasma membrane integrity which showing their best in winter. However, the quality of semen collected and frozen throughout the year was acceptable for AI. Experiment V, aimed to evaluate the distribution of frozen-thawed spermatozoa after laparoscopic artificial insemination (LAI) and to compare the effect of sperm numbers and deposition sites (unilateral and bilateral sites) on pregnancy rate. The post-thaw spermatozoa were stained with CellTrackerTM Green CMFDA (CT-Green) or CellTrackerTM Red CMPTX (CT-Red), and then were deposited into the left and right uterine horns, respectively. The sperm were collected separately from each side of uterus and oviduct after an ovariohysterectomy. Spermatozoa deposited into only one uterine horn distributed to both ipsilateral and contralateral sides. The LAI was performed in 60 does using different numbers of spermatozoa (60 and 120 x106 sperm) and deposition sites (unilateral or bilateral). The pregnancy rates were not significantly different in does deposited with different doses or insemination sites (P>0.05). The unilateral insemination with 60 x106 sperm was sufficient for LAI in goat. Experiment VI, aimed to use LAI and embryo transfer (ET) to produce crossbred black-colored goat. In Experiment 1, LAI with frozen-thawed semen of black buck (Australian Melaan) was performed in 75% Saanen crossbred does (white color, n=70). The total numbers of 68 kids were born from 50 does. The skin colors of kids born were black (10.29%), white (39.71%) and other colors (50%). In Experiment 2, two cross-breeding programs were tested including program I: frozen semen of Australian Melaan inseminated to Black Bengal female (n=7) and program II: frozen semen of Black Bengal inseminated to 50% Australian Melaan (n=7). Thirty embryos at 4-8 cell stage (day 3) were surgically collected and transferred into 30 recipients resulting 30% of pregnancy. Nine kids born from both protocols were black in color with 2.56±0.95 kg birth weights. LAI and ET can be successfully combined to produce and sustain the genetic potential encoding the black-skin color. In conclusion, by an improvement of semen freezing via the semen extender, a study of seasonal effect on semen collection and freezabitity as well as the practical techniques for LAI and ET, can be applied to a goat industry for genetic improvement with an acceptable pregnancy rate.