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Comparison of fertility, regular returns-to-estrus, and calving interval between Ovsynch and CO-synch + CIDR protocols in dairy cows
Theriogenology 82 (2014) 910–914
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Comparison of fertility, regular returns-to-estrus, and calving interval between Ovsynch and CO-synch þ CIDR protocols in dairy cows C. Azevedo a, I. Maia a, N. Canada b, *, J. Simões c a b c
Medicina de Produção Leiteira Veterinária Lda. (MPLVET), Tocha, Portugal Department of Veterinary Clinics, Abel Salazar Institute of Biomedical Sciences, Rua Jorge Viterbo Ferreira, Porto, Portugal Department of Veterinary Sciences, University of Trás-os-Montes e Alto Douro, Vila Real, Portugal
a r t i c l e i n f o
a b s t r a c t
Article history: Received 19 December 2013 Received in revised form 29 May 2014 Accepted 3 July 2014
The main aims of the present study were to compare the pregnancy rate (PR), regular returns-to-estrus, and calving interval of a CO-Synch þ controlled internal drug release (CIDR) device, commonly used to synchronize ovulations in beef cows, with the classical Ovsynch protocol in high-producing dairy cows. Holstein-Friesian cows (n ¼ 128) from six commercial dairy herds, 40 days postpartum and not previously inseminated, were randomly assigned to one of two treatments. Cows submitted to Ovsynch protocol (group OS as control group; n ¼ 66) received 10 mg of a GnRH analogue 7 days before and 48 hours after 25 mg PGF2a, followed by artificial insemination (AI) 16 hours after the second GnRH administration. Cows submitted to CO-Synch þ CIDR (1.38 g of progesterone) inserted for 7 days beginning at the first GnRH administration (group CoS þ CD; n ¼ 62) had the second administration of GnRH concurrent with AI, 64 hours after CIDR removal/PGF2a administration. Nonpregnant cows with return-to-estrus between 18 and 24 days after first AI were reinseminated (second AI). Logistic regressions were used to analyze PR and returns-to-estrus. No effect of group or herd was observed in PR at first timed AI. However, the sum of cows pregnant at first AI and nonpregnant cows with regular returns-to-estrus and the total PR (first þ second AI) were influenced by group treatment. Overall, cows of group CoS þ CD (total PR ¼ 56.5%) were 2.1 times more likely to became pregnant after AI and until first regular returns-to-estrus than cows of group OS. The calving interval was lower in group CoS þ CD (425.9 78.8 days; SD) than in group OS (475.3 83.7 days). The CO-Synch þ CIDR protocol was reliable to use in dairy herds and provided reproductive advantages when compared with Ovsynch protocol. Ó 2014 Elsevier Inc. All rights reserved.
Keywords: Artificial insemination CO-Synch þ CIDR Dairy cattle Fertility Ovsynch
1. Introduction The reproductive performance of lactating dairy cattle remains a major factor affecting herd’s profitability. The increase in dry matter intake and milk production per cow in the last decades has been concurrent with a dramatic
* Corresponding author. Tel.: þ351 252 660 406; fax: þ351 252 661 780. E-mail address: [email protected] (N. Canada). 0093-691X/$ – see front matter Ó 2014 Elsevier Inc. All rights reserved. http://dx.doi.org/10.1016/j.theriogenology.2014.07.006
decline in fertility in dairy commercial herds [1,2]. A negative relation was observed between circulating progesterone (P4) concentrations and dry matter intake in lactating dairy cows [3]. In intensively managed dairy herds with high-producing dairy cows, a negative association between high milk production and expression of behavioral estrus was also observed [1,4,5], showing the importance of timed artificial insemination (TAI) as part of synchronization strategies, with or without estrus detection. The new generation of reproductive management tools has reached recent developments to eliminate
C. Azevedo et al. / Theriogenology 82 (2014) 910–914
detection of estrus and increase cattle handling efficiency. These tools, based on sequential PGF2a and GnRH administrations, or its analogues, before TAI, are focusing on both corpus luteum and follicle control [6], resulting in economical and practical protocols to synchronize ovulation [7]. Recent strategies include P4 intravaginal supplementation as part of classic synchronization protocols for TAI, such as Ovsynch, and several studies have demonstrated increased pregnancy rate (PR), up to 40% at first AI in dairy cows [8–10]. This controlled internal drug release (CIDR) device can also synchronize the return-to-estrus of nonpregnant cows [11]. In beef cows, the addition of an intravaginal P4 device to a CO-Synch protocol, that is, the TAI is concurrent with second GnRH administration, also resulted in improved PR, greater than 50%, but with less one intervention in animals [12–14]. The CO-Synch þ CIDR protocol is, now, the primary TAI protocol recommended for use in beef cows by the American Beef Reproduction Task Force [7]. In fact, the COSynch þ CIDR protocol, by demanding one less handling of cows compared with Ovsynch, increases labor efficiency and reduces the risk of noncompliance that can achieve 30% [15]. To the best of our knowledge, the CO-Synch þ CIDR is a protocol experimented in dairy herds. Initially, DeJarnette and Marshall [16] obtained only a PR of 22% in adult dairy cows. However, PR at first AI greater than 30% was observed by Bartolome et al. [17] and Chebel et al. [18] in highproducing cows. In dairy heifers, the PR also reached 50% using this protocol [19,20]. However, more studies are necessary to evaluate the PR after CO-Synch þ CIDR in adult high-producing cows and subsequent reproductive activity of nonpregnant animals after TAI. The main aims of this study were to determine the PR, regular returns-to-estrus, and calving interval in dairy cows after CO-Synch þ CIDR and to compare these reproductive performance with Ovsynch protocol in commercial herds. 2. Materials and methods 2.1. Population study This experiment was conducted between April 2009 and March 2010 in six commercial dairy herds in the northwest of Portugal. Each farm milked 60 to 70 HolsteinFriesian cows, twice daily. The mean 305-day lactation yields of these farms varied approximately from 9000 to 11,000 kg according the data obtained from the Portuguese National Association for the Improvement of Dairy Cattle. Dairy cattle were housed in free stall barns throughout the year and fed total mixed rations balanced for milk production. In all farms, the total mixed rations consisting of 61.8 0.9% (SD; n ¼ 6) forage and 38.2 0.9% concentrate in dry matter (hay, corn silage, and concentrate for most of the year, as well as grass silage during spring and summer). These total mixed rations were offered for ad libitum consumption considering a daily milk production between 26 and 28 kg. Each cow was also individually supplemented several times daily with concentrate feed according their milk production when greater than 28 kg.
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According the Sea and Atmospheric Portuguese Institute, the monthly mean of maxima surface air temperature was 20.9 5.8 C (n ¼ 12) at herds location throughout the studied period. In all farms, the ventilation was provided by lateral and top gaps. The local air temperature was also regulated by automatic fans when greater than 24 C during the warm days. 2.2. Study design Cows (n ¼ 128), 40 days postpartum and not previously inseminated, without apparent reproductive tract diseases at clinical and transrectal ultrasonography examination, were systematic randomly assigned to receive one of two treatments, at each monthly veterinary visit during the study period in all six herds. After sequential assignment, all selected cows were treated in the next days or weeks according the reproduction management plan of each farm. Cows assigned for group OS (control group; n ¼ 66) received the Ovsynch protocol. Two administrations (im) of 10 mg of a GnRH analogue (2.5 mL Receptal, MSD Animal Health, Germany) were given 7 days before and 48 hours after a 25-mg (im) PGF2a administration (5 mL Dinolitic; Pfizer Animal Health, Belgium, or 2.5 mL Estrumate; MSD Animal Health) followed by TAI 16 hours after second GnRH administration. Cows assigned for group CoS þ CD (CO-Synch þ CIDR group; n ¼ 62) received the Ovsynch protocol plus a CIDR containing 1.38 g of P4 (EAZIBREED CIDR Cattle insert, 1.38 g P4; Pfizer Animal Health, New York, NY, USA) inserted for 7 days, beginning at the first GnRH administration. The CIDR insert was removed at PGF2a administration time. The second GnRH administration was concurrent with TAI, 64 hours after PGF2a administration and CIDR removal. The AI was performed by local dairy cooperative inseminators (n ¼ 3) at farmer request. A weekly rotation route of these experienced inseminators was performed, and Holstein-Friesian semen was chosen by herd manager at each location. A total of 81 sires were used in the six farms during the study period. After first service, estrus detection (estrous behavior) was carried out using visual observation, routinely performed twice in day, in all remaining groups. Cows that returned to estrus 18 to 24 days after first AI, that is, regular returns-to-estrus, were considered nonpregnant at this time and were reinseminated (second AI) according to the AM–PM rule. Others than these first and/or second AI, the nonpregnant cows were also inseminated when natural estrus was detected. Pregnancy status was accessed and confirmed by ultrasonography (linear array 6/8 MHz transrectal transducer, Tringa Linear, Esaote Pie Medical, Genoa, Italy) in all cows after 28 days after TAI and reconfirmed in the next visit. The later embryonic or/and early fetal losses were evaluated by the difference of pregnant cows between these two dates. The PR at first AI was calculated by dividing the number of cows diagnosed pregnant after AI by the number of cows receiving the first AI in each treatment. The PR at second AI was calculated for second service in cows with regular returns-to-estrus. The total PR was calculated by total
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number of cows pregnant until the second AI per total number of cows in each group. The calving-to-(first) AI interval, calving interval, culling rate due to infertility or others problems, and dead or selling of cows were also registered in each farm. The data of 305-day lactation milk yield for each cow (Portuguese Association for the Improvement of Dairy Cattle) were also used. 2.3. Statistical analysis Multiple logistic regression models were used to test the effect of group, farm, season, and inseminator (independent variables) on the PR and regular returns-to-estrus percentage (nominal dependent variables: 1 positive; 0 negative). The effect of group and farm on culling rate was also tested using multiple logistic regression analysis. Main effects and their interactions were considered significant for Wald statistic at P value less than 0.05. The season variable was defined as spring (March, April, and May), summer (June, July, and August), autumn (September, October, and November), and winter (December, January, and February). Mean differences (mean SD) of parity, age, milk production at 305-day lactation, calving-to-first AI interval, and calving interval between groups and farms or their interactions were tested by ANOVA and the Tukey test or by Multivariate Analysis of Variance, respectively. Although the normal population distribution for each continuous variable was previously evaluated by Kolmogorov–Smirnov test, the Van Der Waerden test was also used. The JMP 7 software statistical package was used [21].
Table 2 Mean (SD) age, parity, milk production, and reproductive parameters of cows according to each group. Parameter
Group OS
CoS þ CD
No. of cows Mean age (mo) Mean parity Mean 305-d milk yield (kg)a Calving at first AI interval (d) Culling rate (infertility) Total culling rate (%) Calving interval (d)b
66 60.7 36.0 2.7 1.5 10077 1192 137.0 46.9 9.1% (6/66) 18.2% (12/66) 475.3 83.7c
62 54.5 21.0 2.5 1.5 10082 1256 124.5 36.9 1.6% (1/62) 14.5% (9/62) 425.9 78.8d
Abbreviation: AI, artificial insemination. a 305-day milk yield (kg): average of 305-day milk production levels determined from official records. Group OS (n ¼ 60); group CoS þ CD (n ¼ 57). b Group OS (n ¼ 43); group CoS þ CD (n ¼ 49). Differences of total number of cows between the total culling rate and calving interval were due to animal selling (before parturition) in each farm. c,d Values with different superscripts in the same row differ significantly (P < 0.01).
The calving interval was lower (P < 0.01, Table 2) in group CoS þ CD than in group OS. However, the calving first AI interval and others several parameters, such as culling rates, mean parity, and age and milk production of cows integrated in the present study, were not influenced (P > 0.05) by group and farm or interactions between them. The milk production was only influenced by farm, from 9338 928 kg to 10821 1572 kg (P < 0.01). The global later embryonic and/or early fetal loss observed between days 40.7 10.5 and 62.4 11.8 was 2.9% (2/70).
3. Results
4. Discussion
The PR at first AI, PR at second AI, and the returns-toestrus percentage were not influenced (P > 0.05) by group, farm, inseminator, and season factors. However, a group effect (P < 0.05) on the sum of number of pregnant cows at the first AI and number of nonpregnant cows with regular returns-to-estrus and on the total PR was observed (Table 1). No effect (P > 0.05) of all others independent variables or their interactions was observed. The odds ratio (OR) of group CoS þ CD was 2.17 (95% interval confidence: 1.01–4.64) and 2.13 (95% interval confidence: 1.05–4.31) for the sum of number of pregnant cows at the first AI and number of nonpregnant cows with regular returns-to-estrus and for total PR, respectively.
The PR at first AI was not negatively affected by group, suggesting that the CO-Synch þ CIDR protocol was efficient to synchronize ovulations in high-producing dairy cows. In fact, despite the small animal sample, the PR at first AI obtained in the present study, using this hormonal protocol was 40.3% and is in accordance with the results of Bartolome et al. [17] and Chebel et al. [18] who observed a PR of 35.1% and 33.6%, respectively. However, in those studies, TAI was performed 72 hours after the prostaglandin administration and all the cows were presynchronized. In our study, the TAI performed at 64 hours suggested that this time should be also considered in dairy cows when the CIDR is used. In beef heifers, a PR increase of 10.3% was observed by Kasimanickam et al. [14] moving the time of AI from 72 to 56 hours after CIDR removal in a 5-day COSynch þ CIDR protocol. It is consensual that the AI should take place 10 to 12 hours before ovulation in cows, which occurs with some variability 70 to 75 hours after the PGF2a administration/CIDR removal [22]. However, the second GnRH administration can decrease this variability after CIDR removal in order to use TAI. In fact, a mean interval of 77.5 9.0 hours between the CIDR removal and ovulation after an Ovsynch þ CIDR protocol, but without the second GnRH administration, was observed in Holstein heifers by Hittinger et al. [23], suggesting the use of GnRH at AI time or 12 to 16 hours before TAI, in order to reduce this variation.
Table 1 Descriptive statistic of PR and returns-to-estrus and group effect after logistic regression analysis. Parameter
PR at first AI Regular returns-to-estrus PR at first AI þ regular returns-to-estrus PR at second AI Total PR
Group
P value
OS
CoS þ CD
33.3% (22/66) 38.6% (17/44) 59.1% (39/66)
40.3% (25/62) 59.4% (22/37) 75.8% (47/62)
0.43 0.26 0.04
17.6% (3/17) 37.9% (25/66)
45.5% (10/22) 56.5% (35/62)
0.04
Abbreviations: AI, artificial insemination; PR, pregnancy rate.
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Like CO-Synch þ CIDR, the PR at first AI was also acceptable using Ovsynch (33.3%) protocols in these Portuguese dairy farms. In others studies, this PR ranged from 19.8% to 33.0% after Ovsynch protocol and from 29.0% to 45.1% when the CIDR was inserted (Ovsynch þ CIDR protocol) [8–10,24,25]. Although in several of these studies an improvement of PR by the addition of the CIDR insert was observed [8–10,24], this increase does not always occur [25]. The CIDR use can also improve PR due to decreased pregnancy losses during high environment temperature in warm months [26]. Although during our study the mean monthly maxima surface air temperature in the region was 20.9 5.8 C, all farms had active ventilation system working greater than 24 C and only 2.9% (2/70) of embryonic/fetal losses of all pregnancies were observed. Additionally, the PR at first AI, PR at second AI, and total PR were not influenced by season. In the present study, the TAI was performed without estrus detection in both treated groups. In dairy herds with poor estrus detection rate, like the one observed by Rocha et al. [27] in north of Portugal, the hormonal protocols for ovulation synchronization can reduce the calving-to-first AI interval and consequently the calving interval. Additionally, the present CO-Synch protocol þ CIDR, by demanding one less handling of cows (TAI concurrent with GnRH administration) compared with Ovsynch protocols, can increase labor efficiency and reduce the risk of noncompliance, thus achieving better results. In our study, the group CoS þ CD showed a greater global proportion of cows pregnant at the first AI or with regular returns-to-estrus (75.8% vs. 59.1%; P < 0.05) and a higher total PR (56.5% vs. 37.9%; P < 0.05) than in group OS. On the other hand, the positive effect of CIDR in synchronization of returns-to-estrus (34.1% vs. 19.3% of cows in 3 days for CIDR and no CIDR groups, respectively; P < 0.001) had already been observed by Chenault et al. [11] and confirmed by Alnimer [26] during warm months. Bicalho et al. [28] also observed a greater proportion (17.4%) of cows with P4 1 ng/mL (presence of a functional corpus luteum) at Day 58 3 postpartum using a presynchronization protocol with CIDR when compared with control group (30.6%; P < 0.001). More recently, depending on the CIDR use or not, Chebel et al. [18] observed a higher proportion (P < 0.01) of cows with P4 1 ng/mL (94.4 vs. 86.9%) and P4 3.2 ng/mL (81.8 vs. 68.0%) on days 11 to 14 after TAI. The positive effects of CIDR on ovarian and estrous cycle activities were well demonstrated in these last studies and are in consonance with the fact that the cows of group CoS þ CD of our work were 2.2 times more likely (P < 0.05) to present pregnancy or regular returns-to-estrus (if not pregnant) after TAI than cows of group OS (75.8% vs. 59.1%, respectively). However, considering the total PR in our study, cows of group CoS þ CD were also 2.1 times more likely (P < 0.05) to become pregnant than cows of group OS (56.5% vs. 37.9%, respectively). These results could indicate positive effects of CIDR and/ or GnRH second administration delay or a potential synergic effect between P4 priming (CIDR) and GnRH administration at AI time for maintenance of regular estrous
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cycles in nonpregnant cows after first AI and/or in animals with early embryonic death. Besides oocyte maturation and ovulation, the LH surge induces the onset of luteinization of granulosa and thecal cells, even before ovulation [29,30]. On the other hand, when GnRH administration is performed concurrent with TAI, the LH surge, triggered by CIDR removal, may have already occurred [31]. In fact, Siddiqui et al. [32] observed that the GnRH-induced or natural ovulation occurs 27.1 0.3 hours and 34.5 1.5 hours after the LH surge, respectively. The lower mean calving interval observed in group CoS þ CD (426.0 11.3 days) than in group OS (475.3 12.8 days; P < 0.01) was the probable reflex of the group effect on some reproductive parameters analyzed in the present study. This difference, at least more than 30 days, suggest that the CO-Synch þ CIDR can improve the reproductive efficiency in high-producing dairy cows. In fact, the calving-to-first AI interval was similar for both groups and was not affected by interaction with farms effect or by the percentage of culling cows. 4.1. Conclusions In summary, the CO-Synch þ CIDR seems to be a good alternative to Ovsynch protocol for routine use in dairy farms in order to synchronize ovulations and returns-toestrus. Acknowledgments The authors thank the owners of the farms studied in this article for their cooperation, to Carla Oliveira for her technical assistance, to the Portuguese National Association for the Improvement of Dairy Cattle (http://www.bovinfor. pt/) for the data availability of bovine milk production, and to the Sea and Atmospheric Portuguese Institute (www. ipma.pt/) for the local data availability of monthly surface air temperatures. References [1] Cutullic E, Delaby L, Gallard Y, Disenhaus C. Towards a better understanding of the respective effects of milk yield and body condition dynamics on reproduction in Holstein dairy cows. Anim Int J Anim Biosci 2012;6:476–87. [2] Kawashima C, Matsui M, Shimizu T, Kida K, Miyamoto A. Nutritional factors that regulate ovulation of the dominant follicle during the first follicular wave postpartum in high-producing dairy cows. J Reprod Dev 2012;58:10–6. [3] Hutchinson IA, Dewhurst RJ, Evans ACO, Lonergan P, Butler ST. Effect of grass dry matter intake and fat supplementation on progesterone metabolism in lactating dairy cows. Theriogenology 2012;78:878–86. [4] Cutullic E, Delaby L, Causeur D, Michel G, Disenhaus C. Hierarchy of factors affecting behavioural signs used for oestrus detection of Holstein and Normande dairy cows in a seasonal calving system. Anim Reprod Sci 2009;113:22–37. [5] Crowe MA, Williams EJ. Triennial lactation symposium: effects of stress on postpartum reproduction in dairy cows. J Anim Sci 2012; 90:1722–7. [6] Pursley JR, Kosorok MR, Wiltbank MC. Reproductive management of lactating dairy cows using synchronization of ovulation. J Dairy Sci 1997;80:301–6. [7] Lamb GC, Dahlen CR, Larson JE, Marquezini G, Stevenson JS. Control of the estrous cycle to improve fertility for fixed-time artificial insemination in beef cattle: a review. J Anim Sci 2010;88(13 Suppl): E181–92.
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Comparison of fertility, regular returns-to-estrus, and calving interval between Ovsynch and CO-synch þ CIDR protocols in dairy cows C. Azevedo a, I. Maia a, N. Canada b, *, J. Simões c a b c
Medicina de Produção Leiteira Veterinária Lda. (MPLVET), Tocha, Portugal Department of Veterinary Clinics, Abel Salazar Institute of Biomedical Sciences, Rua Jorge Viterbo Ferreira, Porto, Portugal Department of Veterinary Sciences, University of Trás-os-Montes e Alto Douro, Vila Real, Portugal
a r t i c l e i n f o
a b s t r a c t
Article history: Received 19 December 2013 Received in revised form 29 May 2014 Accepted 3 July 2014
The main aims of the present study were to compare the pregnancy rate (PR), regular returns-to-estrus, and calving interval of a CO-Synch þ controlled internal drug release (CIDR) device, commonly used to synchronize ovulations in beef cows, with the classical Ovsynch protocol in high-producing dairy cows. Holstein-Friesian cows (n ¼ 128) from six commercial dairy herds, 40 days postpartum and not previously inseminated, were randomly assigned to one of two treatments. Cows submitted to Ovsynch protocol (group OS as control group; n ¼ 66) received 10 mg of a GnRH analogue 7 days before and 48 hours after 25 mg PGF2a, followed by artificial insemination (AI) 16 hours after the second GnRH administration. Cows submitted to CO-Synch þ CIDR (1.38 g of progesterone) inserted for 7 days beginning at the first GnRH administration (group CoS þ CD; n ¼ 62) had the second administration of GnRH concurrent with AI, 64 hours after CIDR removal/PGF2a administration. Nonpregnant cows with return-to-estrus between 18 and 24 days after first AI were reinseminated (second AI). Logistic regressions were used to analyze PR and returns-to-estrus. No effect of group or herd was observed in PR at first timed AI. However, the sum of cows pregnant at first AI and nonpregnant cows with regular returns-to-estrus and the total PR (first þ second AI) were influenced by group treatment. Overall, cows of group CoS þ CD (total PR ¼ 56.5%) were 2.1 times more likely to became pregnant after AI and until first regular returns-to-estrus than cows of group OS. The calving interval was lower in group CoS þ CD (425.9 78.8 days; SD) than in group OS (475.3 83.7 days). The CO-Synch þ CIDR protocol was reliable to use in dairy herds and provided reproductive advantages when compared with Ovsynch protocol. Ó 2014 Elsevier Inc. All rights reserved.
Keywords: Artificial insemination CO-Synch þ CIDR Dairy cattle Fertility Ovsynch
1. Introduction The reproductive performance of lactating dairy cattle remains a major factor affecting herd’s profitability. The increase in dry matter intake and milk production per cow in the last decades has been concurrent with a dramatic
* Corresponding author. Tel.: þ351 252 660 406; fax: þ351 252 661 780. E-mail address: [email protected] (N. Canada). 0093-691X/$ – see front matter Ó 2014 Elsevier Inc. All rights reserved. http://dx.doi.org/10.1016/j.theriogenology.2014.07.006
decline in fertility in dairy commercial herds [1,2]. A negative relation was observed between circulating progesterone (P4) concentrations and dry matter intake in lactating dairy cows [3]. In intensively managed dairy herds with high-producing dairy cows, a negative association between high milk production and expression of behavioral estrus was also observed [1,4,5], showing the importance of timed artificial insemination (TAI) as part of synchronization strategies, with or without estrus detection. The new generation of reproductive management tools has reached recent developments to eliminate
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detection of estrus and increase cattle handling efficiency. These tools, based on sequential PGF2a and GnRH administrations, or its analogues, before TAI, are focusing on both corpus luteum and follicle control [6], resulting in economical and practical protocols to synchronize ovulation [7]. Recent strategies include P4 intravaginal supplementation as part of classic synchronization protocols for TAI, such as Ovsynch, and several studies have demonstrated increased pregnancy rate (PR), up to 40% at first AI in dairy cows [8–10]. This controlled internal drug release (CIDR) device can also synchronize the return-to-estrus of nonpregnant cows [11]. In beef cows, the addition of an intravaginal P4 device to a CO-Synch protocol, that is, the TAI is concurrent with second GnRH administration, also resulted in improved PR, greater than 50%, but with less one intervention in animals [12–14]. The CO-Synch þ CIDR protocol is, now, the primary TAI protocol recommended for use in beef cows by the American Beef Reproduction Task Force [7]. In fact, the COSynch þ CIDR protocol, by demanding one less handling of cows compared with Ovsynch, increases labor efficiency and reduces the risk of noncompliance that can achieve 30% [15]. To the best of our knowledge, the CO-Synch þ CIDR is a protocol experimented in dairy herds. Initially, DeJarnette and Marshall [16] obtained only a PR of 22% in adult dairy cows. However, PR at first AI greater than 30% was observed by Bartolome et al. [17] and Chebel et al. [18] in highproducing cows. In dairy heifers, the PR also reached 50% using this protocol [19,20]. However, more studies are necessary to evaluate the PR after CO-Synch þ CIDR in adult high-producing cows and subsequent reproductive activity of nonpregnant animals after TAI. The main aims of this study were to determine the PR, regular returns-to-estrus, and calving interval in dairy cows after CO-Synch þ CIDR and to compare these reproductive performance with Ovsynch protocol in commercial herds. 2. Materials and methods 2.1. Population study This experiment was conducted between April 2009 and March 2010 in six commercial dairy herds in the northwest of Portugal. Each farm milked 60 to 70 HolsteinFriesian cows, twice daily. The mean 305-day lactation yields of these farms varied approximately from 9000 to 11,000 kg according the data obtained from the Portuguese National Association for the Improvement of Dairy Cattle. Dairy cattle were housed in free stall barns throughout the year and fed total mixed rations balanced for milk production. In all farms, the total mixed rations consisting of 61.8 0.9% (SD; n ¼ 6) forage and 38.2 0.9% concentrate in dry matter (hay, corn silage, and concentrate for most of the year, as well as grass silage during spring and summer). These total mixed rations were offered for ad libitum consumption considering a daily milk production between 26 and 28 kg. Each cow was also individually supplemented several times daily with concentrate feed according their milk production when greater than 28 kg.
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According the Sea and Atmospheric Portuguese Institute, the monthly mean of maxima surface air temperature was 20.9 5.8 C (n ¼ 12) at herds location throughout the studied period. In all farms, the ventilation was provided by lateral and top gaps. The local air temperature was also regulated by automatic fans when greater than 24 C during the warm days. 2.2. Study design Cows (n ¼ 128), 40 days postpartum and not previously inseminated, without apparent reproductive tract diseases at clinical and transrectal ultrasonography examination, were systematic randomly assigned to receive one of two treatments, at each monthly veterinary visit during the study period in all six herds. After sequential assignment, all selected cows were treated in the next days or weeks according the reproduction management plan of each farm. Cows assigned for group OS (control group; n ¼ 66) received the Ovsynch protocol. Two administrations (im) of 10 mg of a GnRH analogue (2.5 mL Receptal, MSD Animal Health, Germany) were given 7 days before and 48 hours after a 25-mg (im) PGF2a administration (5 mL Dinolitic; Pfizer Animal Health, Belgium, or 2.5 mL Estrumate; MSD Animal Health) followed by TAI 16 hours after second GnRH administration. Cows assigned for group CoS þ CD (CO-Synch þ CIDR group; n ¼ 62) received the Ovsynch protocol plus a CIDR containing 1.38 g of P4 (EAZIBREED CIDR Cattle insert, 1.38 g P4; Pfizer Animal Health, New York, NY, USA) inserted for 7 days, beginning at the first GnRH administration. The CIDR insert was removed at PGF2a administration time. The second GnRH administration was concurrent with TAI, 64 hours after PGF2a administration and CIDR removal. The AI was performed by local dairy cooperative inseminators (n ¼ 3) at farmer request. A weekly rotation route of these experienced inseminators was performed, and Holstein-Friesian semen was chosen by herd manager at each location. A total of 81 sires were used in the six farms during the study period. After first service, estrus detection (estrous behavior) was carried out using visual observation, routinely performed twice in day, in all remaining groups. Cows that returned to estrus 18 to 24 days after first AI, that is, regular returns-to-estrus, were considered nonpregnant at this time and were reinseminated (second AI) according to the AM–PM rule. Others than these first and/or second AI, the nonpregnant cows were also inseminated when natural estrus was detected. Pregnancy status was accessed and confirmed by ultrasonography (linear array 6/8 MHz transrectal transducer, Tringa Linear, Esaote Pie Medical, Genoa, Italy) in all cows after 28 days after TAI and reconfirmed in the next visit. The later embryonic or/and early fetal losses were evaluated by the difference of pregnant cows between these two dates. The PR at first AI was calculated by dividing the number of cows diagnosed pregnant after AI by the number of cows receiving the first AI in each treatment. The PR at second AI was calculated for second service in cows with regular returns-to-estrus. The total PR was calculated by total
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number of cows pregnant until the second AI per total number of cows in each group. The calving-to-(first) AI interval, calving interval, culling rate due to infertility or others problems, and dead or selling of cows were also registered in each farm. The data of 305-day lactation milk yield for each cow (Portuguese Association for the Improvement of Dairy Cattle) were also used. 2.3. Statistical analysis Multiple logistic regression models were used to test the effect of group, farm, season, and inseminator (independent variables) on the PR and regular returns-to-estrus percentage (nominal dependent variables: 1 positive; 0 negative). The effect of group and farm on culling rate was also tested using multiple logistic regression analysis. Main effects and their interactions were considered significant for Wald statistic at P value less than 0.05. The season variable was defined as spring (March, April, and May), summer (June, July, and August), autumn (September, October, and November), and winter (December, January, and February). Mean differences (mean SD) of parity, age, milk production at 305-day lactation, calving-to-first AI interval, and calving interval between groups and farms or their interactions were tested by ANOVA and the Tukey test or by Multivariate Analysis of Variance, respectively. Although the normal population distribution for each continuous variable was previously evaluated by Kolmogorov–Smirnov test, the Van Der Waerden test was also used. The JMP 7 software statistical package was used [21].
Table 2 Mean (SD) age, parity, milk production, and reproductive parameters of cows according to each group. Parameter
Group OS
CoS þ CD
No. of cows Mean age (mo) Mean parity Mean 305-d milk yield (kg)a Calving at first AI interval (d) Culling rate (infertility) Total culling rate (%) Calving interval (d)b
66 60.7 36.0 2.7 1.5 10077 1192 137.0 46.9 9.1% (6/66) 18.2% (12/66) 475.3 83.7c
62 54.5 21.0 2.5 1.5 10082 1256 124.5 36.9 1.6% (1/62) 14.5% (9/62) 425.9 78.8d
Abbreviation: AI, artificial insemination. a 305-day milk yield (kg): average of 305-day milk production levels determined from official records. Group OS (n ¼ 60); group CoS þ CD (n ¼ 57). b Group OS (n ¼ 43); group CoS þ CD (n ¼ 49). Differences of total number of cows between the total culling rate and calving interval were due to animal selling (before parturition) in each farm. c,d Values with different superscripts in the same row differ significantly (P < 0.01).
The calving interval was lower (P < 0.01, Table 2) in group CoS þ CD than in group OS. However, the calving first AI interval and others several parameters, such as culling rates, mean parity, and age and milk production of cows integrated in the present study, were not influenced (P > 0.05) by group and farm or interactions between them. The milk production was only influenced by farm, from 9338 928 kg to 10821 1572 kg (P < 0.01). The global later embryonic and/or early fetal loss observed between days 40.7 10.5 and 62.4 11.8 was 2.9% (2/70).
3. Results
4. Discussion
The PR at first AI, PR at second AI, and the returns-toestrus percentage were not influenced (P > 0.05) by group, farm, inseminator, and season factors. However, a group effect (P < 0.05) on the sum of number of pregnant cows at the first AI and number of nonpregnant cows with regular returns-to-estrus and on the total PR was observed (Table 1). No effect (P > 0.05) of all others independent variables or their interactions was observed. The odds ratio (OR) of group CoS þ CD was 2.17 (95% interval confidence: 1.01–4.64) and 2.13 (95% interval confidence: 1.05–4.31) for the sum of number of pregnant cows at the first AI and number of nonpregnant cows with regular returns-to-estrus and for total PR, respectively.
The PR at first AI was not negatively affected by group, suggesting that the CO-Synch þ CIDR protocol was efficient to synchronize ovulations in high-producing dairy cows. In fact, despite the small animal sample, the PR at first AI obtained in the present study, using this hormonal protocol was 40.3% and is in accordance with the results of Bartolome et al. [17] and Chebel et al. [18] who observed a PR of 35.1% and 33.6%, respectively. However, in those studies, TAI was performed 72 hours after the prostaglandin administration and all the cows were presynchronized. In our study, the TAI performed at 64 hours suggested that this time should be also considered in dairy cows when the CIDR is used. In beef heifers, a PR increase of 10.3% was observed by Kasimanickam et al. [14] moving the time of AI from 72 to 56 hours after CIDR removal in a 5-day COSynch þ CIDR protocol. It is consensual that the AI should take place 10 to 12 hours before ovulation in cows, which occurs with some variability 70 to 75 hours after the PGF2a administration/CIDR removal [22]. However, the second GnRH administration can decrease this variability after CIDR removal in order to use TAI. In fact, a mean interval of 77.5 9.0 hours between the CIDR removal and ovulation after an Ovsynch þ CIDR protocol, but without the second GnRH administration, was observed in Holstein heifers by Hittinger et al. [23], suggesting the use of GnRH at AI time or 12 to 16 hours before TAI, in order to reduce this variation.
Table 1 Descriptive statistic of PR and returns-to-estrus and group effect after logistic regression analysis. Parameter
PR at first AI Regular returns-to-estrus PR at first AI þ regular returns-to-estrus PR at second AI Total PR
Group
P value
OS
CoS þ CD
33.3% (22/66) 38.6% (17/44) 59.1% (39/66)
40.3% (25/62) 59.4% (22/37) 75.8% (47/62)
0.43 0.26 0.04
17.6% (3/17) 37.9% (25/66)
45.5% (10/22) 56.5% (35/62)
0.04
Abbreviations: AI, artificial insemination; PR, pregnancy rate.
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Like CO-Synch þ CIDR, the PR at first AI was also acceptable using Ovsynch (33.3%) protocols in these Portuguese dairy farms. In others studies, this PR ranged from 19.8% to 33.0% after Ovsynch protocol and from 29.0% to 45.1% when the CIDR was inserted (Ovsynch þ CIDR protocol) [8–10,24,25]. Although in several of these studies an improvement of PR by the addition of the CIDR insert was observed [8–10,24], this increase does not always occur [25]. The CIDR use can also improve PR due to decreased pregnancy losses during high environment temperature in warm months [26]. Although during our study the mean monthly maxima surface air temperature in the region was 20.9 5.8 C, all farms had active ventilation system working greater than 24 C and only 2.9% (2/70) of embryonic/fetal losses of all pregnancies were observed. Additionally, the PR at first AI, PR at second AI, and total PR were not influenced by season. In the present study, the TAI was performed without estrus detection in both treated groups. In dairy herds with poor estrus detection rate, like the one observed by Rocha et al. [27] in north of Portugal, the hormonal protocols for ovulation synchronization can reduce the calving-to-first AI interval and consequently the calving interval. Additionally, the present CO-Synch protocol þ CIDR, by demanding one less handling of cows (TAI concurrent with GnRH administration) compared with Ovsynch protocols, can increase labor efficiency and reduce the risk of noncompliance, thus achieving better results. In our study, the group CoS þ CD showed a greater global proportion of cows pregnant at the first AI or with regular returns-to-estrus (75.8% vs. 59.1%; P < 0.05) and a higher total PR (56.5% vs. 37.9%; P < 0.05) than in group OS. On the other hand, the positive effect of CIDR in synchronization of returns-to-estrus (34.1% vs. 19.3% of cows in 3 days for CIDR and no CIDR groups, respectively; P < 0.001) had already been observed by Chenault et al. [11] and confirmed by Alnimer [26] during warm months. Bicalho et al. [28] also observed a greater proportion (17.4%) of cows with P4 1 ng/mL (presence of a functional corpus luteum) at Day 58 3 postpartum using a presynchronization protocol with CIDR when compared with control group (30.6%; P < 0.001). More recently, depending on the CIDR use or not, Chebel et al. [18] observed a higher proportion (P < 0.01) of cows with P4 1 ng/mL (94.4 vs. 86.9%) and P4 3.2 ng/mL (81.8 vs. 68.0%) on days 11 to 14 after TAI. The positive effects of CIDR on ovarian and estrous cycle activities were well demonstrated in these last studies and are in consonance with the fact that the cows of group CoS þ CD of our work were 2.2 times more likely (P < 0.05) to present pregnancy or regular returns-to-estrus (if not pregnant) after TAI than cows of group OS (75.8% vs. 59.1%, respectively). However, considering the total PR in our study, cows of group CoS þ CD were also 2.1 times more likely (P < 0.05) to become pregnant than cows of group OS (56.5% vs. 37.9%, respectively). These results could indicate positive effects of CIDR and/ or GnRH second administration delay or a potential synergic effect between P4 priming (CIDR) and GnRH administration at AI time for maintenance of regular estrous
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cycles in nonpregnant cows after first AI and/or in animals with early embryonic death. Besides oocyte maturation and ovulation, the LH surge induces the onset of luteinization of granulosa and thecal cells, even before ovulation [29,30]. On the other hand, when GnRH administration is performed concurrent with TAI, the LH surge, triggered by CIDR removal, may have already occurred [31]. In fact, Siddiqui et al. [32] observed that the GnRH-induced or natural ovulation occurs 27.1 0.3 hours and 34.5 1.5 hours after the LH surge, respectively. The lower mean calving interval observed in group CoS þ CD (426.0 11.3 days) than in group OS (475.3 12.8 days; P < 0.01) was the probable reflex of the group effect on some reproductive parameters analyzed in the present study. This difference, at least more than 30 days, suggest that the CO-Synch þ CIDR can improve the reproductive efficiency in high-producing dairy cows. In fact, the calving-to-first AI interval was similar for both groups and was not affected by interaction with farms effect or by the percentage of culling cows. 4.1. Conclusions In summary, the CO-Synch þ CIDR seems to be a good alternative to Ovsynch protocol for routine use in dairy farms in order to synchronize ovulations and returns-toestrus. Acknowledgments The authors thank the owners of the farms studied in this article for their cooperation, to Carla Oliveira for her technical assistance, to the Portuguese National Association for the Improvement of Dairy Cattle (http://www.bovinfor. pt/) for the data availability of bovine milk production, and to the Sea and Atmospheric Portuguese Institute (www. ipma.pt/) for the local data availability of monthly surface air temperatures. References [1] Cutullic E, Delaby L, Gallard Y, Disenhaus C. Towards a better understanding of the respective effects of milk yield and body condition dynamics on reproduction in Holstein dairy cows. Anim Int J Anim Biosci 2012;6:476–87. [2] Kawashima C, Matsui M, Shimizu T, Kida K, Miyamoto A. Nutritional factors that regulate ovulation of the dominant follicle during the first follicular wave postpartum in high-producing dairy cows. J Reprod Dev 2012;58:10–6. [3] Hutchinson IA, Dewhurst RJ, Evans ACO, Lonergan P, Butler ST. Effect of grass dry matter intake and fat supplementation on progesterone metabolism in lactating dairy cows. Theriogenology 2012;78:878–86. [4] Cutullic E, Delaby L, Causeur D, Michel G, Disenhaus C. Hierarchy of factors affecting behavioural signs used for oestrus detection of Holstein and Normande dairy cows in a seasonal calving system. Anim Reprod Sci 2009;113:22–37. [5] Crowe MA, Williams EJ. Triennial lactation symposium: effects of stress on postpartum reproduction in dairy cows. J Anim Sci 2012; 90:1722–7. [6] Pursley JR, Kosorok MR, Wiltbank MC. Reproductive management of lactating dairy cows using synchronization of ovulation. J Dairy Sci 1997;80:301–6. [7] Lamb GC, Dahlen CR, Larson JE, Marquezini G, Stevenson JS. Control of the estrous cycle to improve fertility for fixed-time artificial insemination in beef cattle: a review. J Anim Sci 2010;88(13 Suppl): E181–92.
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