In P4/E2-based protocols, it was observed that eCG applications had positive effects on fertility in cows undergoing timed artificial insemination (Sá Filho, Ayres, et al., 2010; Dorneles Tortorella et al., 2013; Núñez-Olivera et al., 2014; Bilbao et al., 2015). It was thought that treatment with eCG would increase gonadotropin support in follicle development and thus increase the size of the dominant follicle and the possibility of ovulation, thereby increasing fertility in grazing purebred or crossbred beef and dairy cattle (Prata et al., 2017). It has been reported that eCG application had no significant effect on fertility in cyclic or anestrus Nelore cows (Barreiros et al., 2014). It has also been reported that Nelore heifers treated with eCG had more ovarian response and pregnancy than heifers not treated with eCG (Sá Filho, Torres-Júnior, et al., 2010) Similarly, in Nelore cows in the early postpartum period undergoing progesterone-based timed artificial insemination with appropriate body condition, it was observed that eCG application and removal of the calf from the mother did not affect the pregnancy rate (Pinheiro et al., 2009). In addition, eCG treatment in the first synchronized estrus improved reproductive performance in the short term; however, it did not affect it after subsequent inseminations (Rowe et al., 2019). In another study, no significant differences were found between the follicular dynamics of cyclic and anestrus groups treated with TCR (56 hours breastfeeding) or eCG, despite the differences in the cyclicity of the animals. Follicular diameter, follicular growth rate, corpus luteum size, and P4 concentrations of anestrous cows treated with eCG or TCR were similar (Barreiros et al., 2014).
In the present study, the effect of eCG injection on pregnancy rates after intravaginal application with progesterone, GnRH, and PGF2α-based protocols and timed artificial insemination was evaluated in Brown Swiss cows. With the removal of the progesterone-releasing intravaginal device and application of eCG, development of the dominant follicle was accelerated, and ovulation was stimulated with the LH peak. According to the present study results, pregnancy rates were 10% higher in cows treated with eCG; however, no statistical differences were found. The higher pregnancy rate was associated with possible corpus luteum lysis with PGF2α injection one day before PRID®Delta removal and accelerated follicle development with eCG application. The research was conducted on cows that did not show any symptoms of estrus within two months postpartum. Cows exited the early postpartum period when the postpartum negative energy balance was established. In the early postpartum period, cows may have insufficient gonadotropin support, and therefore fertility may be positively affected in response to eCG treatment. After cows are out of negative energy balance in the early postpartum period, additional gonadotropin supplementation may not be required. Therefore positive effects of eCG may not be observed in cows after 70 days postpartum (Prata et al., 2017).
It has been reported that eCG application during the removal of the vaginal device in cows with anestrus increased the ovulation rate and improved luteal functions (Núñez-Olivera et al., 2014) and eGG administration can reduce differences in ovulation timing (Cavalieri et al., 1997). However, another study reported that ovulation rates did not increase after eCG (Pulley et al., 2013). The eCG treatment yields a larger corpus luteum and higher progesterone concentrations in cows that ovulate after synchronization (Núñez-Olivera et al., 2014). In the present study, it was seen that intravaginal devices containing progesterone could increase serum progesterone concentration in cows without estrus symptoms. For the cyclic activity to develop healthily and the follicular activity in the ovary to continue, the genital organs must be under the influence of progesterone for a certain period. The progesterone value was lower in cows treated with eCG during artificial insemination. The low level of progesterone leads to the peak of LH release and increases the likelihood of ovulation. This low concentration of progesterone may have caused the difference in pregnancy rates.
Long-term progesterone treatment did not increase the pregnancy rate in cows with high BCS; however, the combination of long-term progesterone and eCG treatment increased the pregnancy rate per preovulatory follicle and artificial insemination in cows with low BCS (Bilbao et al., 2015). The effect of eCG applications on fertility in lactating Holstein cows was limited to cows with low BCS. The eCG treatment improved ovulation rates or corpus luteum development, especially in cows with low VKS, and it has been reported that this may lead to increased fertility in cows. It has been observed that cows with a BCS <2.75 have increased follicle growth and fertility with eCG treatment in those with insufficient LH support (Souza et al., 2009). In the present study, no differences were found in pregnancy rates of anestrus cows with or without eCG injection, with a VKS ≥ 2.75 or a VKS < 2.75. Different factors affect the anestrus mechanism in cows other than VKS. In lactating dairy cows under the Presynch-Ovsynch program, eCG treatment at a dose of 400 IU three days before artificial insemination at the first service did not increase the size of the largest follicle, follicle growth rate, or pregnancy rate 48 hours after application. It was thought that the dose and timing of eCG treatment could be delayed to affect the maturation period of the preovulatory follicle (Pulley et al., 2013). One study has reported that eCG application did not increase the ovulation rate. However, in another study, it was found that follicle size, corpus luteum volume, serum P4 concentrations, and pregnancy rates were higher when eCG was applied two days before the removal of the intravaginal progesterone device rather than when eCG was applied during the removal of the device in cows undergoing timed artificial insemination. (Dorneles Tortorella et al., 2013). During the application of timed artificial insemination, the diameter of the largest follicle found in the ovaries was larger in heifers treated with eCG than in untreated heifers (Sá Filho, Ayres, et al., 2010). Follicle diameters of anestrous cows and cyclic animals treated with eCG were similar. It has been shown that cows with eCG or anestrus treated by removing calves from suckling for 56 hours showed follicular diameter and a follicular growth rate similar to cyclic cows (Barreiros et al., 2014). The addition of eCG to progesterone and estradiol-based treatments increased the ovulation rate of the dominant follicle in anestrous cows due to the stimulating effect of eCG on growth rate (Núñez-Olivera et al., 2014). The most important result obtained in the present study was that the follicle diameter at the time of artificial insemination was larger in cows that did not show signs of estrus with eCG injection compared to cows that were not injected with eCG. It was observed that this difference in follicle diameter did not affect pregnancy rates, however, it is thought that eCG application increased the growth rate of the follicle in anestrous cows. The larger diameter of the largest follicle in the eCG group can be explained by the fact that eCG increases FSH and LH release for follicle development. The increase in follicular diameter induced by eCG may be related to the positive effect on steroidogenic cells in the ovulatory follicle and the formation of a more competent corpus luteum observed in the treated animals (Núñez-Olivera et al., 2014).
In conclusion, it was determined that progesterone (PRID®Delta)-based GnRH, PGF2α application in addition to eCG injections increased the pregnancy rate numerically in cows without symptoms of estrus. It was also determined that the largest follicle diameter was greater in cows treated with eCG during artificial insemination; however, follicle size did not increase the pregnancy rate.