Livestock Research for Rural Development 28 (9) 2016 Guide for preparation of papers LRRD Newsletter

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Effects of genotype and FSH dose on estrus and ovarian response of Boran and Boran x Holstein Friesian cows in Ethiopia

Tamrat Degefa, Alemayehu Lemma1, Azage Tegegne2 and C R Youngs3

Debre Zeit Agricultural Research Center, Ethiopian Institute of Agricultural Research
1 Addis Ababa University, College of Veterinary Medicine and Agriculture
2 International Livestock Research Institute (ILRI), Addis Ababa, Ethiopia
3 Department of Animal Science, Iowa State University, Ames, Iowa 50011 USA


This study utilized 27 Boran and 20 Boran x Holstein Friesian cows 6-8 years of age. A 7-day CIDR treatment was initiated on Day 1, and cows were treated on Days 4-7 with 200, 250, or 300 IU Pluset® administered at 12-hour intervals in a decreasing dose regimen. Prostaglandin F2α was given one day before CIDR withdrawal, and cows were monitored for 84 hours to determine the onset and duration of estrus. Ovarian response was assessed by counting the number of corpora lutea (CL) via transrectal ultrasonography.

The proportion of cows that exhibited estrus in response to treatment was 76.6% and was higher (P=0.003) for Boran than for Boran x Holstein Friesian crosses. The mean interval from CIDR removal to the onset of estrus was 20.4 ± 1.8 hours; this interval was not affected (P>.10) by genotype but was affected by Pluset® dose with a shorter (P<0.01) interval (10.8 ± 3.3 hr) for cows receiving 250 IU. Duration of estrus was not affected (P>0.10) by genotype but was influenced by dose with a shorter (P<0.009) duration of estrus (16.8 ± 2.1 hr) in cows treated with 250 IU Pluset®. Boran cows had a higher (P<0.03) number of CL (10.1 ± 0.7) than Boran x Holstein Friesian crosses (7.2 ± 0.9). Results indicate that, under conditions of our study in Ethiopia, Boran cows have better ovarian response to exogenous gonadotropins than Boran x Holstein Friesian crosses.

Key words: cattle, corpus luteum, superovulation


It has been reported previously that Bos indicus cattle are less fertile and have lower levels of milk production than Bos taurus breeds; however, they are better adapted to harsh environmental conditions which makes them more likely to reproduce successfully in the tropics (Negussie et al 1999). Fundamental biological differences in reproductive function between Bos taurus and Bos indicus cattle have been reported with respect to age at puberty, estrous cycle patterns, estrous behavior, acquisition of ovulatory capacity, ovarian structures and reproductive hormone production (Sartori et al 2010). The main physiological differences include higher circulating concentrations of ovarian steroid hormones (e.g., estradiol and progesterone), larger population of small diameter ovarian follicles, smaller size of the dominant follicle at deviation, and greater sensitivity of ovarian follicles to gonadotropins in Bos indicus compared with Bos taurus cattle (Sartori et al 2010).

Multiple ovulation and embryo transfer (MOET) technology is widely used globally in the cattle industry, with more than 575,000 in vivo derived embryos transferred annually (Perry 2015). Superovulatory treatment of donor cows enables production of a higher number of transferable quality embryos and higher probability of producing pregnancies (Mapletoft et al 2002). Variability in the response to superovulatory treatments is a major impediment to further adoption of this technology, and different breeds of cattle seemingly respond differently to similar superovulation protocols. It is well accepted that the dose of exogenous FSH required for superovulation of Bos indicus cattle is less than that for Bos taurus cattle (Barati et al 2006), and Bos indicus breeds require 25-30% less FSH than Bos taurus breeds (Kanitz et al 2002). Barati et al (2006) reported that a superovulatory regime of 250 mg Folltropin-V (considered a moderate to low dose for Bos taurus cattle) caused ovarian overstimulation and a high incidence of anovulatory follicles in Sastani cattle (a native Bos indicus breed of Iran).

In spite of extensive research conducted on the reproductive physiology of Bos indicus breeds in many areas of the tropics (Abeygunawardena and Dematawewa 2004; Bastos et al 2010; Carvalho et al 2008; Eler et al 2006; Forni and Albuquerque 2005; Gimenes et al 2009; Mollo et al 2007; Nogueira 2004; Pereira et al 2007), studies on bovine reproductive physiology in other parts of the tropics seem to be merely adaptations of reproductive management strategies developed for Bos taurus breeds. An increase in knowledge of the physiological differences between Bos taurus and Bos indicus breeds that have not yet been extensively studied would be useful for the development of specific reproductive management protocols and/or strategies to maximize production efficiency of different Bos indicus breeds of cattle raised in a tropical or semi-tropical environment. One such breed that has not yet been fully characterized is the Boran, an indigenous breed of Ethiopia which is also found in other east African nations. The Boran breed is one of five indigenous cattle breeds of Ethiopia listed as endangered by FAO (Philipsson 1992).

The objective of this study was to assess the effect of dose of Pluset® (a product that contains follicle stimulating hormone and luteinizing hormone) on the estrous and ovarian responses of purebred Boran and Boran x Holstein Friesian crossbred cows raised under semi-tropical conditions in Ethiopia.

Materials and methods

The experiment was carried out at the Debre Zeit Agricultural Research Center (DZARC) of the Ethiopian Agricultural Research Institute dairy cattle improvement farm, located about 45 km east of Addis Ababa (8°46′13.57"N, 38°59′50.45"E; 1900 meters above sea level). The average annual temperature for the last five years was 18.5°C with an average annual rainfall of 757.05 mm (DZARC Agro-meteorology 2015).

A total of 47 cows between 6 and 8 years of age with an average body condition score of 7.0 (range 6.0 to 8.0) on a scale of 1 to 9 (1= emaciated to 9=obese) were selected as donor cows after passing a thorough gynecological examination. All cows were managed under similar housing and health management conditions. Both groups of cows were fed tef (Eragrostis tef) straw and grass hay as a basal diet that was supplemented with commercially prepared concentrate feed (wheat bran, wheat middling, corn, Noug cakes [oil seed cake = Guizotia abyssinica], and mineral salts). Cows were provided water ad libitum.

The cows were blocked by genotype for superovulatory treatments; Group I consisted of 27 purebred Boran, and Group II consisted of 20 Boran x Holstein Friesian crossbreds. Cows in each group received a CIDR (1.38 gm progesterone, DEC International NZ Ltd) for seven days. On Day 4 of CIDR treatment, cows were allotted within genotype to one of three superovulatory treatments: 200 IU, 250 IU, or 300 IU Pluset® (Barcelona, Spain). The Pluset® was given as a series of eight intramuscular injections administered each morning and afternoon in a decreasing dose regimen. The 300 IU regimen was: 60, 60, 45, 45, 30, 30, 15, and 15 IU. The 250 IU regimen was: 50, 50, 35, 35, 25, 25, 15, and 15 IU. The 200 IU regimen was: 40, 40, 30, 30 20, 20, 10, and 10 IU. All cows received 2 ml of i.m. cloprostenol (Estrumate®, Schering-Plough BPK) on the sixth day of CIDR treatment, and the CIDR was withdrawn on seventh day at 6 PM. For 84 hours after CIDR removal all cows were monitored a minimum of 6 times per day to determine the onset and duration of estrus. The number of corpora lutea (CL) present on each ovary was determined on Day 7 via transrectal ultrasonography (CTS3300, Italy, 5 MHz linear array probe). Superovulation was considered successful if a cow exhibited estrus and possessed more than one CL at the time ovaries were examined.

For the binary response variable of estrus/no estrus, data were analyzed via Chi-square. Data on the onset and duration of estrus, as well as the number of CL, were analyzed via analysis of variance (IBM® SPSS® Statistics 20). Mean separations were performed using the least significant difference test of the generalized linear model procedure.

Because this was a pilot experiment on the feasibility of inducing multiple ovulations with a commercially available exogenous gonadotropin preparation (Pluset®), embryo recovery data are not reported.


The proportion of cows that exhibited estrus was 76.6%, and the incidence of estrus was higher (χ2 = 9.057; p=0.003) in Boran (92.6%) than in Boran x Holstein Friesian crossbred cows (55.0%).

For those cows that exhibited estrus, the mean (± SEM) time interval from CIDR withdrawal to the onset of estrus was 20.4 ± 1.8 hours, and this interval was not affected (P>.10) by genotype (Table 1). However, cows treated with 250 IU exhibited a shorter (P<0.01) interval from CIDR removal to estrus (10.8 ± 3.3 hrs) than cows treated with either 200 IU or 300 IU Pluset®.

The mean duration of estrus was 22.9 ± 1.1 hours and was not affected (P>.10) by genotype. However, the duration of estrus was shorter (P<0.005) for cows treated with 250 IU Pluset® compared with those treated with 200 IU or 300 IU (Table 1).

Table 1. Time interval from CIDR withdrawal to onset of estrus and duration of estrus in Boran and Boran x Holstein Friesian crosses treated with different doses of exogenous Pluset®



dose [IU]

Mean (±SEM)
interval to estrus (hrs)

Mean (±SEM)
duration of estrus (hrs)




18.6 ± 2.1

21.8 ± 1.4

Boran x Holstein


19.8 ± 2.9

22.8 ± 1.9


Pluset® dose



22.6 ± 3.1a

26.1 ± 1.9a



10.8 ± 3.3b

16.8 ± 2.1b



24.2 ± 2.8a

24.0 ± 1.8a


Genotype x Dose




23.2 ± 2.9

24.9 ± 1.7



10.8 ± 4.7

15.5 ± 2.9



21.8 ± 2.9

24.4 ± 1.8


Boran x Holstein



22.0 ± 5.5

27.3 ± 3.4



10.8 ± 4.7

17.0 ± 2.9



26.5 ± 4.7

24.0 ± 2.9

ab Means with unlike superscripts in column sub-sets are different at P<0.01

The mean (±SEM) number of CL counted ultrasonographically was 8.6 ± 0.7. A summary of the ovarian response to treatment with Pluset® is illustrated in Table 2. Boran cows had a greater (P<0.05) ovarian response to superovulation than Boran X Holstein Friesian cows. Cows treated with 250 IU Pluset® produced a greater (P<0.05) number of CL than those treated with 200 IU; cows treated with 300 IU were intermediate (Table 2). The maximum number of CL counted on any individual female was 17, and this was detected in a Boran cow stimulated with 250 IU.

Table 2. The number of corpora lutea (CL) in Boran and Boran x Holstein Friesian crosses treated
with different doses of exogenous Pluset®



Ultrasound CL count‡
Mean (±SEM)




10.1 ± 0.7a

Boran x Holstein


7.2 ± 0.9b


Pluset® Dose (IU)



7.0 ± 1.0d



10.6 ± 1.0c



8.3 ± 0.9cd


Genotype x Dose

Boran 200


8.0 ± 0.9

Boran 250


13.8 ± 1.5

Boran 300


8.6 ± 0.9

Boran x Holstein 200


6.0 ± 1.7

Boran x Holstein 250


7.5 ± 1.5

Boran x Holstein 300


8.0 ± 1.5

‡ Denotes the number of corpora lutea (CL) counted during ultrasonographic ovarian examinations
ab Means within a column sub-set with unlike superscripts are different at P<0.02
cd Means within a column sub-set with unlike superscripts are different at P<0.05


The proportion of cows exhibiting estrus in response to exogenous Pluset® treatment in the present study (76.6%) was higher than that previously reported for superovulated cows of other African breeds such as the Cheurfa in Algeria (46.6%; Ferrouk et al 2008) and the Oulmes-Zaer in Morocco (53.0% and 48.7%; Elaidi et al 1996a and Elaidi et al 1996b, respectively).

The mean number of CL observed in the present study with Boran (10.1) was higher than that reported for the Sistani breed in Iran (8.7 CL; Barati et al 2006), the Cheurfa breed in Algeria (7.5 CL; Ferrouk et al 2008), the Baoulé breed of central Africa (6.9 CL: Bianchi et al 1986; 6.5 CL: Chicoteau 1989), and the N’dama breed of west Africa (5.1 CL: Diop et al 1994; 4.5 CL: Jordt et al 1986). Unlike the report by Ferrouk et al (2008) in the Cheurfa breed, considerable variation was observed between Boran and Boran x Holstein crosses in their superovulatory response. The difference in response to superovulation is related to dose and genotype effects, and agrees with the previous report by Kanitz et al (2002).

It was interesting to note that the number of CL (10.1 CL) found in Boran cows superovulated in the present study with 200-300 IU was higher than a previous report from Ethiopia with Boran cattle superovulated with much higher doses (500 IU or 1000 IU) of the same commercially available superovulatory product (7.8 CL; Tegegne et al 1994). Similarly, Elaidi et al (1996a) reported fewer CLs in Oulmes-Zaer cows (7.2 CL) in one study, but more CL in Oulmes-Zaer cows (10.5 CL) in another study (Elaidi et al 1996b). A more recent study on cows of the Curraleiro Pé-duro breed of northeastern and central Brazil revealed a greater number of CL (12.3 CL; Teixeira et al 2013) than what we observed in our study.

In the present study the ovarian response of purebred Boran cows to a relatively low dose of exogenous gonadotropins was quite acceptable and gives strong encouragement for future studies of the production of in vivo derived preimplantation embryos from this breed. Although other Bos indicus breeds of cattle (Kamphaeng Saen: Nilchuen et al 2012; Nelore: Carvalho 2004) produce a higher number of CL in response to superovulation than the Boran, finding the optimal dose of exogenous FSH for each breed is important. Barati et al (2006), for example, reported that a superovulatory dose of 250 mg Folltropin-V caused ovarian overstimulation and a high incidence of unovulated ovarian follicles in Sistani cattle, a native Bos indicus breed of Iran.

Previous studies by Kanitz et al (2002), Murphy et al (1998), and Escouflaire et al (1989) documented an important relationship between the dose of exogenous FSH administered for superovulation and the resultant number of ovulations and number of transferrable quality embryos recovered. The number of ovulations increases with increasing doses of exogenous FSH up to an optimal dose level, and then falls off as the dose of exogenous FSH increases above optimal. Kanitz et al (2002) proposed that a dose of exogenous FSH above the optimum can lead to a disruption of the normal ovulatory mechanisms, and there is ample biological evidence that ovarian overstimulation can lead to: a) ovaries that are too large for the infundibulum to catch all ovulated oocytes, b) induction of endogenous inhibin production, reducing the levels of endogenous FSH produced by the female [thus reducing superovulatory response], and c) alteration of gamete transport due to elevated concentrations of estradiol in the bloodstream of superovulated cows.

Our findings with purebred Boran and crossbred Boran X Holstein cows are in strong agreement with previous findings pertaining to the dose of exogenous FSH and its relationship with superovulatory response. Moreover, our study further confirms results of previous studies which indicated that Zebu cattle (Bos indicus) require less exogenous FSH than Bos taurus breeds to achieve optimal superovulatory response. Further research on factors impacting the response to superovulation (e.g., nutritional status, reproductive history, age, season, breed, ovarian status at the time of treatment, repeated superstimulation; Mapletoft 2006) will be needed for the Boran breed. It is important to preserve the Boran breed in a pure state as this breed is well adapted to the harsh environmental conditions of Ethiopia and other parts of east Africa.


Conflict of interest

The authors declare that there is no conflict of interest for this study.


Use of animals in this study was reviewed and approved by the Iowa State University Committee on Animal Care. The authors gratefully acknowledge financial support of this study by the United States Department of Agriculture (USDA) Foreign Agriculture Service (FAS) Borlaug Fellowship Program, the United States Agency for International Development (USAID) Borlaug LEAP (Leadership Enhancement in Agriculture Program) Program, the East African Agricultural Productivity Project (EAAPP) of Ethiopian Institute of Agricultural Research, Iowa State University, and the USDA multi-state research project W-3171 “Germ Cell and Embryo Development and Manipulation for the Improvement of Livestock”. The technical assistance of Dr. Gebre Meskel Mamo, Mr. Jeiu Jemal, Ms. Asnakech Funga, and Mr. Biniam Abebe is also gratefully acknowledged.


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Received 2 March 2015; Accepted 30 May 2016; Published 1 September 2016

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