Timed Artificial Insemination: Pregnancy Rates in Sokoto Gudali Cattle Treated with Prostaglandin F2 Alpha at a Private Dairy Farm in Nigeria.

Pregnancy rates (PR) resulting from timed artificial insemination (TAI) with frozen semen were investigated in 192 Sokoto Gudali (SG) cattle following estrus synchronization under 11- and 12- day treatment intervals with prostaglandin F2 alpha (Estrumate®). Two-point inseminations at 60 h and 72 h post Estrumate® treatments were used. All cattle were randomly allocated to 2 groups i.e. A and B, such that each sub-group of A1, A2, B1 and B2 had sixteen (16) cows and thirty two (32) heifers. Furthermore, cattle in Groups A and B were administered double Estrumate® injections at 11- and 12-day intervals, respectively. Cattle in sub-groups A1 and B1 were inseminated at 60 h post second Estrumate® injection while A2 and B2 were inseminated at 72 h. Pregnancy was diagnosed using the 60-day non return to estrus and confirmed via rectal palpation at 90 days post insemination. The results showed the following PR: 33%, 83%, 33% and 33% for cattle in sub-groups A1, A2, B1 and B2, respectively. It is concluded therefore that timed artificial insemination at 72 h in SG cattle treated with double Estrumate® injections at 11-day interval produced the best pregnancy rate. These findings raises hope on the future application of TAI in breeding the SG cattle with prostaglandin F2 alpha as single agent of estrus synchronization.


Timed
Artificial Insemination (TAI) programmes describe estrous synchronization protocols wherein estrus detection prior to insemination are not necessarily required. The success of TAI programmes depend much on the ability of the synchronization protocol/agent to achieve ovulation synchrony. At the present, many protocols, involving several agents, have been developed that perfectly fit into TAI. While some reports (Garcia and Salaheddine, 2001;Martinez et al., 2005) attest to the efficacy of Estradiol 17ß in producing high degree of ovulation synchrony in heifers, Colazo and Mapletoft (2014) have described Estradiol-based TAI protocols as obsolete. Many other TAI-relevant protocols such as CoSynch, SelectSynch and Presynchronization are modifications of the OvSynch protocol that was first described by Pursley et al. (1995). The Ovsynch protocol is composed of a first gonadotropin (GnRH) injection on day zero (d 0), followed by prostaglandin F2 alpha (d 7) and a second GnRH (d 9), after which cows are covered with semen between 8-18 hours thereafter (Pursley et al., 1997). The advantages of Ovsynch and its modifications include high submission and pregnancy rates (PR) in cows as well as overall improvement in reproductive performance (El-Zarkouny et al., 2004;Pursley et al., 1997). Prostaglandin F2 alpha (PGF) is involved in nearly all TAI protocols mentioned above. It functions to regress the corpus lutem (CL) (Day and Geary, 2005;Giordano et al., 2013), the source of progesterone, and thus, removing its inhibitory effects on the hypothalamo-pituitary axis. Subsequently, endogeneous GnRH is released for the regulation of the estrous cycle. An interesting fact is that PGF e.g. Estrumate ® is a single agent capable of synchronizing estrus. Its effectiveness have been shown in caprine (Leigh et al., 2010), bovine (Venkata et al., 2013), ovine (Omontese et al., 2014) and lately swine (Alimi et al., 2017). Few of PGF limitations however are its inability to regulate follicular waves (Mihm and Bleach, 2003), leading to poor ovulation synchrony, cows and heifers without a sensitive CL are also not responsive (Kastelic et al., 1990). The CL following recent ovulation (0-4 days) is refractory to PGF, while 5-16 d post ovulation is senstive and by 17-21 d is already regressing (Wenzinger and Bleul, 2012). These variability results in wide variations in responses of cows and heifers to PGF treatment and hence poor PR, especially with the protocol involving single injection. The protocol involving double injections 10-14 days apart produces better result compared to the earlier, and depending on considerations of cost of hormone and labour involved, either options may be favoured. Some researchers have carried out trials with single and double PGFinjection protocol for prediction of best time for insemination. One study reported 61.4%, 45.7% and 46.9% PR to TAI at 72 and 96 hrs (h) post single PGF treatment in zebu cows at three different farms (Rekwot et al., 1999), while another (Voh Jr. et al., 2000) reported 54.3%, 45.7%, 41.4% and 40.0% to TAI at 60, 72, 84 and 96 h post second PGF treatments, respectively. That two fixed-time inseminations at 72 and 96 h were carried out (Rekwot et al., 1999) presupposes that PR ought to be higher than where single insemination was done at either period in agreement with the findings of Helmuth et al. (2005). Also, that PR at 60 h was better than at later hours (Voh Jr. et al., 2000) would alert inseminators that though peak estrus response occurs 2-3 days following PGF treatment, about 12 h to the end of that 9window[ could be the optimum time to obtain highest PR. Since the Sokoto Gudali (SG) cattle continues to occupy a critical position in the Nigerian economy, where also PGF is the most accessible estrus synchronization agent, these claims require more investigation in order to ascertain the optimum insemination time. This study therefore investigated PR in SG cattle subjected to two-point insemination at 60 and 72 h following 11-and 12-day treatment intervals with Estrumate ® .

Materials and Methods
Study site: The study was carried out at a private-owned dairy station established about eight years ago and located in the Ibarapa zone, Oyo State, Nigeria. The station houses a little over 500 cattle heads (all females) of the Bokolooji (Sokoto Gudali) breed.

Study animals and management:
One hundred and ninety two (192) Sokoto Gudali cattle consisting of 64 cows (1-3 parity, 50-60 days post calving) and 128 heifers (175-240 kg) were used for the study. The animals spent most of their time in their pen where they were fed with silage and crushed elephant grass. They were allowed to graze in fenced paddocks for 3-5 hours daily during which their pens were cleaned.

Study design:
The animals were randomly allocated to sub-groups A1, A2, B1 and B2 such that each group comprised of sixteen (16) cows and thirty two (32) heifers. The double PGF protocol was used for estrus synchronization. Animals in A1 and A2 were administered (i/m) 500 µg Estrumate ® (Schering-Plough Animal Health Corp, Germany) at 11 days interval while those in B1 and B2 were administered same doses at 12 days interval. Insemination was carried out by a single experienced inseminator via the recto-cervical/vaginal method (ABS, 2008), using thawed (36 o C) frozen semen from Holstein-Friesian bulls packaged in straws which were previously stored at -196 o C. The semen straws were obtained from the International Fertilization Develpment Center (IFDC). Animals in A1 and B1 were inseminated at 60 h post second Estrumate ® injection while those in A2 and B2 were similarly inseminated at 72 h. The 60-day non return to estrus (Nematolla et al., 2013), was used to investigate cows that were pregnant.
Pregnancies were thereafter confirmed through rectal palpation of the fetus at d 90 post insemination .

Data Collection and Analysis:
The pregnancy rate (PR) was calculated by dividing the number of animals confirmed pregnant via rectal palpation by the number of animals inseminated in the sub-groups (Nematolla et al., 2013). Data were summarized using descriptive statistics (Elston and Johnson, 2008).

Guide on Research Conduct:
The study was conducted under strict adherence to the principles of the care and use of farm animals in research, teaching and testing of the Canadian Council on Animal Use.

Results and Discussion
The results are shown in Tables 1 and 2. Table 1 shows that under 11-day PGF treatment interval, the pregnancy rates (PR) of 33% and 83% were obtained for sokoto Gudali (SG) cattle inseminated at 60 h (A1) and 72 h (A2) post double PGF injections, respectively. Our results clearly indicate that a singletimed artificial insemination at 72 h following prostaglandin-PGF (Estrumate ® ) synchronization in a double injection protocol of 11 days interval gave the highest pregnancy rate (PR). In this study, the PR for Sokoto Gudali (SG) cattle in other groups i.e. 11-day PGF interval 60 h insemination, as well as 60 h and 72 h inseminations under 12-day PGF interval were same and ridiculously low compared to that at 72 h 11-day interval. It is not unlikely that the low PR in these sub-groups could have arisen from several other factors, not excluding poor synchronization rate. These results suggest that for SG cattle, best PR may be obtained with double injections of Estrumate ® given at 11-day interval with insemination carried out at 72 h post treatment. The present finding is not in total agreement with the report of a study conducted in northern Nigeria which found that 60 h after two injections of PGF at 11 days interval was the optimum time for insemination in the White Fulani cattle (Voh Jr. et al., 2000). Although, that study agreed with the present one in terms of treatment interval of 11 days, it disagreed only on the time of insemination. The position of these authors cannot be contested at this level but it will suffice to reiterate that the present study recorded only 33% PR under conditions that the earlier report claimed was best. The present study also shows that insemination around the end of the window for peak estrus response following PGF (Estrumate ® ) treatment, and not 12 h earlier yielded highest PR. While the present authors opined that many factors such as fertility of semen and inseminator efficiency could be responsible for this disparity, it is hoped that breed factor as earlier reported by Mai et al. (2014), did not play a role in the differences observed. In a similar study , Voh Jr. (1996) reported a PR of 74.1% in control cattle that were inseminated 12 h after detected estrus. Our current finding of 83% PR is undoubtedly better than 74.1%, suggesting an advantage in terms of efficiency of PGF synchronization over spontaneous estrus, this is aside the rigours and failures associated with estrus detection. The current findings also appeared to be in consonance, though with a caveat, with the report that in heifers, single insemination at 72 h or 80 h after treatment with PGF gives normal fertility compared with insemination at 48 h or 60 h (Roch, 1977). Our finding with PR at 72 h insemination in the 11-day treatment interval group is in line with this report, albeit, cattle in group B2 had low PR. This observation may further strengthen the importance of PGF treatment interval in cattle synchronization programmes in that a little difference in treatment interval may result in huge losses with regards to PR as shown in this study with group B2. The present finding of 83% PR is also suggestive of an improvement over two-fixed time inseminations at 60 and 72 h following a single administration of PGF in which the highest PR obtained in three farms was 61.4% (Rekwot et al., 1999). Although, It has not been clearly ascertained if double inseminations really has better chances of producing pregnancies more than single inseminations. While some authors (Helmuth et al., 2005) suspected it does, others (Stevenson et al., 1990) have found no benefit in double, over single inseminations. Our findings in the present study has however shown that single insemination at 72 h post PGF in an 11-day treatment interval has produced a very high pregnancy rate of 83% in Sokoto Gudali cattle. This result is better than previous ones reported and therefore suggest that the present protocol has the potential to increase the reproductive efficiency of cattle at breeding stations.

Conclusions
It is concluded therefore that timed artificial insemination at 72 h following double prostaglandin (Estrumate ® ) injections at 11-day treatment interval produced the highest pregnancy rate in Sokoto Gudali cattle.