Freezing of Stallion Semen: In Vitro Evaluation of Motility and Acrosin Activity in Sperm Cells Cryopreserved Using Different Semen Extenders.

The work described here aimed to verify the efficiency of different extenders for cryopreservation of equine semen using sperm motility and acrosin activity as spermatic parameters. The semen was fractioned into two equal parts and resuspended in an 11% lactose solution in a 1:1 proportion, where it remained for 20 minutes at room temperature. The semen was centrifuged at 600 g for 10 minutes, and after the second centrifugation, each pellet received the freezing extender (Merck or Zorlesco) and was loaded into 4 mL straws. Each straw was placed in liquid nitrogen vapor steam for 15 minutes and further immersion in liquid nitrogen at -196°C for long-term storage. After thawing, semen samples were initially evaluated for sperm motility, both total and progressive, and acrosin activity. Moreover, semen was incubated at 37°C and further assessed at 60 and 120 minutes in a thermoresistance test (TRT) for sperm motility and acrosin activity. Immediately after thawing, both progressive and total motility, and acrosin activity were lower (p < 0.05) in thawed semen than in fresh semen. During the TRT, total sperm motility and acrosin activity after 60 minutes were lower (p < 0.05) than those obtained after thawing. Similarly, total sperm motility and acrosin activity were lower (p < 0.05) after 120 minutes than at 60 minutes of the TRT. The analysis of motility and acrosin activity allowed the conclusion that both extenders have a similar capacity to preserve the integrity of sperm cells subject to freezing and thawing.

Conceptus loss in Santa Inês ewes carrying twin pregnancies by natural mating or embryo transfer.

Commercial application of reproductive biotechnologies such as multiple ovulation and embryo transfer depends on its overall efficiency. Sheep embryo transfer is gradually gaining wider adoption, but pregnancy rates after embryo transfer remain lower than those derived from natural mating for most breeds. The work was aimed to evaluate embryonic and fetal losses in Santa Inês ewes carrying twin pregnancies by natural mating or embryo transfer. Ewes were subjected to synchronized natural mating by ram effect or used as recipients for embryo transfer. Ewes diagnosed as carrying twin pregnancies at day 25 were used in the experiment (n = 42). Conceptus viability was monitored by ultrasonography on days 30, 35, 40, 45, 50, and 55 after conception. Conceptus loss was similar (P > 0.05) within natural mating 11/42 (26.19%) and embryo transfer 14/42 (33.34%). However, overall embryonic loss (80.0%) was greater (P < 0.05) than fetal loss (20.0%), with no difference within groups The results allow the conclusion that conceptus loss after embryo transfer is similar to natural mating and occurs predominantly during the embryonic stages.

Ultrasonographic fetal sex identification in pregnant sheep derived from natural mating and embryo transfer.

The objective of this study was to identify the migration period of the genital tubercle and the period of visualization of external genital structures in fetuses of the Dorper breed of sheep derived from natural mating and from fresh, frozen and vitrified embryo transfer. Transrectal ultrasound was performed using a double-frequency linear transducer (6.0 and 8.0 MHz) to monitor 130 ewe fetuses distributed in the four treatments regarding embryo origin. The accuracy of the ultrasound was 100% in this experiment. The fetuses originated from controlled natural mating (NM) and from fresh (FrE), frozen (FE) and vitrified (VE) embryo transfer, with embryos collected 7 days after breeding. Migration of the genital tubercle occurred earlier (P<0.05) in NM (42.21+/-2.86 days) than in FrE (43.98+/-3.00 days), FE (44.97+/-1.83 days) and VE (44.58+/-1.97 days). Visualization of the scrotal bag, prepuce and vulva occurred, respectively, earlier (P<0.05) in NM (45.22+/-1.25, 45.95+/-1.53 and 45.01+/-1.03 days) than in FrE (48.91+/-1.92, 48.52+/-1.41 and 47.41+/-1.41 days), FE (49.97+/-1.08, 49.18+/-2.00 and 47.64+/-1.82 days) and VE (50.12+/-1.66, 49.27+/-1.61 and 47.93+/-1.92 days). The results show that fetal sexing can be accomplished from the 50th day onward in fetuses produced by natural mating and from the 55th day onward in fetuses derived from fresh, frozen and vitrified embryos. It can also be concluded that real-time ultrasonography is a reliable tool for fetal sex determination in sheep taking into account both the location of the genital tubercle and the identification of external genital structures.

Migration time of the genital tubercle in caprine and ovine fetuses: comparison between breeds, sexes and species.

The aim of this work was to determine the ideal moment to sex goat and sheep fetuses, to compare the average time of genital tubercle (GT) migration between sexes, breeds and species, and to evaluate the accuracy of fetal sexing between sexes. A total of 317 fetuses of 219 pregnant females were monitored at 24-hour interval, from days 30 to 60 of pregnancy in ewes, and from days 40 to 60 in goats. Examinations were performed using transrectal ultrasound equipped with a linear transducer of double frequency. Fetuses were identified as male when the GT was next to the umbilical cord and female when the GT was next to the tail. The average time of GT migration in ewes (41.3 +/- 3.1 days) was shorter (P < 0.05) than in goats (47.2 +/- 2.3 days)? In goats, the average time of GT migration of Saanen fetuses was later (P < 0.05) than in fetuses of other breeds, with no difference in the average time of GT migration between male (46.9 +/- 2.2) and female fetuses (47.4 +/- 2.4). In ewes, the average time of GT migration did not differ (P > 0.05) among breeds and sexes. In goat and sheep, no difference was noticed in the accuracy of fetal sexing between males and females (P > 0.05). The results show that fetal sexing in ewes must be done earlier than in goats, fetal sexing in Saanen goats must be performed later, and fetal sex does not influence the time of GT migration in either of the two species.