Paula Alvares Lunardelli

Developmental block and programmed cell death in Bos indicus embryos: effects of protein supplementation source and developmental kinetics.

The aims of this study were to determine if the protein source of the medium influences zebu embryo development and if developmental kinetics, developmental block and programmed cell death are related. The culture medium was supplemented with either fetal calf serum or bovine serum albumin. The embryos were classified as Fast (n = 1,235) or Slow (n = 485) based on the time required to reach the fourth cell cycle (48 h and 90 h post insemination - hpi -, respectively). The Slow group was further separated into two groups: those presenting exactly 4 cells at 48 hpi (Slow/4 cells) and those that reached the fourth cell cycle at 90 hpi (Slow). Blastocyst quality, DNA fragmentation, mitochondrial membrane potential and signs of apoptosis or necrosis were evaluated. The Slow group had higher incidence of developmental block than the Fast group. The embryos supplemented with fetal calf serum had lower quality. DNA fragmentation and mitochondrial membrane potential were absent in embryos at 48 hpi but present at 90 hpi. Early signs of apoptosis were more frequent in the Slow and Slow/4 cell groups than in the Fast group. We concluded that fetal calf serum reduces blastocyst development and quality, but the mechanism appears to be independent of DNA fragmentation. The apoptotic cells detected at 48 hpi reveal a possible mechanism of programmed cell death activation prior to genome activation. The apoptotic cells observed in the slow-developing embryos suggested a relationship between programmed cell death and embryonic developmental kinetics in zebu in vitro-produced embryos.

Resynchronization of estrous cycle with eCG and temporary calf removal in lactating Bos indicus cows

The aim of this study was to compare four methods of estrus resynchronization performed 23 days after timed artificial insemination (TAI) plus estrus observation in Bos indicus cows. Eight hundred fourteen lactating Nelore cows were submitted to TAI and then randomly assigned to one of the five following treatments: R23 (resynchronization without eCG), R23/200 (resynchronization with 200 IU of eCG), R23/300 (resynchronization with 300 IU of eCG), R23/TCR (resynchronization with temporary calf removal [TCR]), and a control group, with estrus observation followed by AI (with no resynchronization). Treatment consisted of a progesterone device plus administration of estradiol benzoate on Day 0; on Day 8, the device was removed and cloprostenol was applied, together with estradiol cypionate. Also on Day 8, either eCG was administered or TCR was performed in the resynchronized groups, except for R23. The females were inseminated 48 hours after device removal or TCR (33 days after the first TAI). The control group was kept under estrus observation from 18 to 23 days after the first TAI and was inseminated 12 hours after detection of estrus. The first pregnancy evaluation was performed using ultrasound examination 31 days after the first TAI. After 30 days of the resynchronization, a second pregnancy evaluation was performed and the animals in the R23/300 and R23/TCR groups achieved the highest conception rates, 76.6% and 74.0%, respectively (P < 0.05). There were no differences between the conception rates of the animals in the R23/200 (63.3%), R23 (61.3%), and control (54.3%) groups (P > 0.05). These results suggest that estrus resynchronization at 23 days after TAI can effectively improve the conception rate of lactating Bos indicus cows in a short time period. Furthermore, resynchronization with 300 IU of eCG or with TCR provided the best results.

A new direct transfer protocol for cryopreserved IVF embryos.

The global demand for in vitro-produced (IVP) embryos of determined sex has greatly increased over the last decade. Efficient protocols for the direct transfer of IVP embryos are lacking. This study aimed to compare the pregnancy rates for fresh, vitrified, or frozen/directly transferred IVP dairy cow embryos. Oocytes (n = 3171) recovered by ovum pickup (n = 112) from Girolando (Holstein-Gir) females (n = 36) were selected and submitted to IVM for 24 hours at 38.5 °C with 5% CO2 in air with saturated humidity. In vitro fertilization was performed with the thawed, sexed semen from 5 Holstein bulls. After IVF, presumptive zygotes were denuded and cultured for 7 days under the same IVM and IVF conditions of temperature and humidity, except with 5% CO2 and 5% O2. Grade I blastocysts were randomly assigned for either the transferred fresh, vitrified/thawing, or frozen/directly embryo transfer into previously synchronized recipient females. Conception rates were analyzed by binomial logistic regression, and a probability level of P < 0.05 was considered significant. The conception rates were 51.35 ± 1.87% (133/259) for the fresh embryos, 35.89 ± 3.87% (84/234) for the vitrified embryos, and 40.19 ± 4.65% (125/311) for the frozen directly transferred embryos. These data demonstrate that IVP embryos with sexed semen could be directly transferred into recipient cows with similar conception rates to vitrified embryos. The comparison found that the use of frozen embryos in direct transfer provides easier logistics and a more practical approach for the transfer of IVP embryos on dairy farms.

Effects of progestagen exposure duration on estrus synchronization and conception rates of crossbreed ewes undergoing fixed time artificial insemination.

Synchronization of estrus and ovulation are of paramount importance in modern livestock improvement programs. These methods are critical for assisted reproduction technologies, including artificial insemination and embryo transfer, that can increase productivity. In the current study, subcutaneous implants containing norgestomet were placed for long (14 days), medium (9 days), and short (5 days) periods of time in 70 crossbred ewes undergoing fixed-time artificial insemination. The resulting effects on estrus synchronization and conception rates were subsequently evaluated. Among the synchronized ewes, 85.7% (60/70) underwent estrus over a period of 72 h after progestagen treatment ceased. The shortest mean interval between withdrawal of the device and onset of estrus (34.2 ± 8.9 h) was observed in the G14 days of P4 group (p < 0.05). The conception rate of the G14 days of P4 group was statistically higher than that of the other groups (83.3% vs. 60.9% vs. 47.8%; p < 0.05). In conclusion, 14 days of norgestomet treatment produced higher conception rates and a greater number of pregnancies at the beginning of the breeding season.