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Efficacy of various five-day estrous synchronization protocols in sheep
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Small Ruminant Research journal homepage: www.elsevier.com/locate/smallrumres
Efficacy of various five-day estrous synchronization protocols in sheep C.G. Jackson a , T.L. Neville a , V.R.G. Mercadante b , K.M. Waters b , G.C. Lamb b , C.R. Dahlen a , R.R. Redden a,∗ a b
Department of Animal Sciences, North Dakota State University, NDSU Dept. 7630, P.O. Box 6050, Fargo, ND 58108-6050, USA North Florida Research and Education Center, University of Florida, 3925 HWY 71, Marianna, FL 32446-8091, USA
a r t i c l e
i n f o
Article history: Received 26 February 2014 Accepted 8 April 2014 Available online xxx
Keywords: Controlled internal drug releasing inserts Estrous synchronization Sheep
a b s t r a c t The objective of this study was to determine the effects of ewe estrus synchronization protocols including combinations of controlled internal drug releasing (CIDR) inserts for 5 d, GnRH, and PG on concentrations of progesterone (P4) and reproductive performance of ewes during different breeding seasons. Experiment 1 was conducted during the transition season (August–October) and Experiment 2 was conducted during the anestrous season (April–May). In both experiments ewes received 1 of 4 treatments: (1) untreated (CON); (2) CIDR (0.3 g P4) insert (CT); (3) CIDR and PG at CIDR removal (CTP); or (4) GnRH at CIDR insertion and PG at CIDR removal (CTG). Rams equipped with marking harnesses were introduced at CIDR removal (d 0) and ewes were observed for breeding marks twice daily at 0800 h and 1700 h. In Experiment 1, days to estrus was greater (P ≤ 0.01) in CON (9.5 ± 1.10 d) and CTP (6.5 ± 1.04 d) compared with CT (3.6 ± 1.03 d). No differences (P = 0.27) were detected in pregnancy rate within the first 17 d of the breeding season for CON, CT, CTP, and CTG treatments (37 ± 7.8%, 62 ± 7.5%, 56 ± 8.1%, and 46 ± 7.7%, respectively). Similarly, no differences (P ≥ 0.28) were detected among treatments for lambing rate and prolificacy. Contrasts for all CIDR treatments compared to controls determined that CIDR treated ewes had less days to estrus and lambing (P ≤ 0.05) than control ewes. In Experiment 2, days to estrus was greater (P ≤ 0.02) in CON (6.5 ± 1.05 d) and CTP (5.9 ± 0.88 d) compared with CTG (3.0 ± 0.88 d). No differences were detected (P ≥ 0.23) among treatments for overall percentage of ewes exhibiting estrus. No differences (P = 0.79) in pregnancy rate within the first 17 d of the breeding season were detected among treatments (42 ± 12.4%, 37 ± 11.3%, 35 ± 11.1%, and 50 ± 10.9% for CON, CT, CTP, and CTG, respectively). No differences (P ≥ 0.44) were observed among treatments in overall lambing rate and prolificacy. In conclusion, inserting a CIDR for 5 d prior to ram introduction regardless of the use of PG or GnRH, reduced the interval to estrus compared with untreated ewes but did not impact overall pregnancy rates, lambing rates, or prolificacy. Published by Elsevier B.V.
1. Introduction
∗ Corresponding author. Tel.: +1 701 231 5597; fax: +1 701 231 7590. E-mail address: [email protected] (R.R. Redden).
Reproductive performance is one of the largest determining factors of profitability in today’s sheep industry and will aid in its future success and long-term sustainability. For the U.S. sheep industry to remain competitive in
http://dx.doi.org/10.1016/j.smallrumres.2014.04.004 0921-4488/Published by Elsevier B.V.
Please cite this article in press as: Jackson, C.G., et al., Efficacy of various five-day estrous synchronization protocols in sheep. Small Ruminant Res. (2014), http://dx.doi.org/10.1016/j.smallrumres.2014.04.004
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the market place and to increase year-round demand for lamb, producers could benefit from technology that would effectively synchronize estrus in ewes during the transition and anestrous seasons. Controlled internal drug releasing (CIDR) inserts were approved for use in sheep by the U.S. Food and Drug Administration in 2009 for estrous synchronization and are currently labeled for 5 d use to induce estrus in anestrous ewes. Due to the seasonal nature of estrous cyclicity of most sheep, exogenous hormones can be used to facilitate the induction and synchronization of estrus in the anestrous period (Carlson et al., 1989). This is of particular interest to those who produce lambs consistently throughout the year in an accelerated lambing system. Producers in a traditional spring lambing system can also benefit from exogenous progesterone (P4) and PG use to synchronize estrus during the transition period (when the amount of day light begins to decrease in the northern and southern latitudes and ewes begin to exhibit estrous cyclicity). Due to the seasonal nature of ovine estrous cyclicity, PG is only effective in animals already cycling; therefore, only a portion of ewes may respond to this treatment. To assist in initiating hormonal events necessary to synchronize estrous cycles, such as estrus and ovulation, GnRH may also be incorporated, especially in anestrous ewes (Carlson et al., 1989; Wildeus, 2000). Therefore, our objective was to determine the efficacy of CIDR insertion for 5 d when used alone or in combination with GnRH and PG during both the transition and anestrous seasons. We hypothesized that estrous synchronization with CIDR, GnRH, and/or PG will decrease days to estrus and increase pregnancy, lambing, and prolificacy rates. 2. Materials and methods The North Dakota State University Institutional Animal Care and Use Committee approved all protocols and procedures involving animals used in this research under protocol A11007. Animals were housed at the North Dakota State University Sheep Unit in Fargo, ND. 2.1. Animals and treatments Experiment 1: In August 2010 and 2011, Columbia (n = 38 and 38, respectively) and Hampshire (n = 47 and 53, respectively) ewes were randomly assigned to 1 of 4 treatments: (1) untreated (CON; n = 44); (2) 5 d CIDR insert (EAZI-BREED CIDR Sheep Insert, 0.3 g P4, Zoetis Animal Health, Florham Park, NJ; CT; n = 45); (3) 5 d CIDR and PG (Lutalyse, 10 mg dinoprost tromethamine i.m., Zoetis Animal Health) at CIDR removal (CTP; n = 44); and (4) GnRH (Factrel, 0.025 mg gonadorelin hydrochloride i.m., Zoetis Animal Health) at CIDR insertion and PG at CIDR removal (CTG; n = 43). Treatments were assigned over 3 time periods, 1 wk apart, to ensure an adequate ram to ewe ratio throughout the breeding period. One week prior to treatment application, ewes were moved to paddocks that had not been previously grazed for 1 mo. Upon treatment administration, all ewes were moved to a drylot with continuous access to fresh water and received alfalfa hay (3 kg/ewe) and a 14% CP concentrate ration (1 kg/ewe) daily. Prior to treatment initiation, ewes were isolated from males. Rams equipped with marking harnesses were single sire mated to ewes within treatment group at a ewe to ram ratio ≤25:1. Ewes were exposed to rams at CIDR insert removal (d 0) for a 60-d breeding season. Ewes were observed twice daily for breeding marks at 0800 h and 1700 h starting on d 0. Ewes were determined to be in estrus as indicated by breeding marks (when ≥60% of the rump was colored with wax marker). Experiment 2: In April 2011, during the anestrous period, multiparous and nulliparous Dorset and Katahdin (n = 61 and n = 17, respectively) ewes from the North Dakota State University Sheep Unit in Fargo, ND, were
randomly assigned to 1 of 4 treatments as described previously in Experiment 1: (1) untreated (CON; n = 16); (2) CIDR inserted for 5 d (CT; n = 21); (3) CIDR inserted for 5 d and PG at removal (CTP; n = 20); and (4) GnRH given at CIDR insertion and PG at CIDR removal (CTG; n = 21). Prior to treatment application, ewes were managed as a common group in a dry lot, isolated from all males, and had ad libitum access to alfalfa hay and fresh water. On d 0, 4 rams equipped with marking harnesses were introduced to ewes immediately post CIDR removal with a ewe to ram ratio ≤25:1. Beginning at d 0, ewes were observed for breeding marks twice daily at 0800 h and 1700 h and were determined to be in estrus as indicated by breeding marks. The breeding season lasted for duration of 42 d. 2.2. Reproductive performance data collection For both experiments data collected included days to estrus, days to lambing, percentage of ewes exhibiting estrus (divided the number of ewes marked by a ram by the total number of ewes on study), pregnancy rate (number of ewes lambing as a percentage of ewes in all treatments), lambing rate (number of lambs/ewe exposed), and prolificacy (number of lambs/ewe lambing). The latter 4 measures of interest were further broken down to represent the length of an estrous cycle (first 17 d of the breeding season), or the entire breeding season (overall; from d 0 until ram removal). 2.3. Blood collection and analysis To evaluate concentrations of serum P4 for Experiments 1 and 2, blood samples were collected via jugular venipuncture into 10-mL serum tubes (BD Vacutainer Serum, Becton, Dickinson and Company, Franklin Lakes, NJ) and immediately placed on ice. Samples were centrifuged at 4 ◦ C for 30 min at 1500 × g. Then serum was transferred into plastic 2.0-mL microcentrifuge tubes and frozen at −20 ◦ C until assayed. In Experiment 1, blood samples were collected 1 wk prior to (d-12) and on the day of treatment initiation (d-5) to obtain baseline concentrations of P4. Additional samples were collected at CIDR removal (d 0) and on d 2, 4, 6, 8, 10, and 12 in 2010 and on d 1, 2, 3, 4, 5, 8, 11, 14, 17, and 20 in 2011. In Experiment 2, samples were collected on d-12, -5, 0, 1, 2, 3, 4, 5, 8, 11, 14, 17, and 20 relative to CIDR removal. To detect circulating concentrations of P4, samples were assayed using a solid-phase, no extraction RIA (Coat-a-Count Progesterone, Diagnostic Products Corporation, Los Angeles, CA). The assay kit was validated for ovine serum using an assay volume of 100 L (Hamra et al., 1986; Schneider and Hallford, 1996). Assay was standardized to a curve of 0.1, 0.25, 0.5, 1.0, 2.0, 5.0, 10.0, and 20.0 ng/mL. Assay sensitivity for a 100-L sample was 0.1 ng/mL. Ewes were determined to be cycling if serum P4 concentrations were greater than 1 ng/mL on either or both the d-12 or -5 samples. All samples for a single ewe were analyzed within the same assay and treatments were run in a random order. The intraand inter-assay CVs for Experiment 1 were 5.39% and 3.61%, respectively and were 2.28% and 2.21%, respectively for Experiment 2. 2.4. Statistical analysis The MIXED procedures of SAS 9.2 (SAS Inst., Inc., Cary, NC) were utilized to examine d to detected estrus and d to lambing. The model included the main effects of treatment, breed, age, cycling status, and their respective interactions. The GLIMMIX procedures of SAS were used to analyze percentage of ewes exhibiting estrus, pregnancy rate, lambing rate, and prolificacy. The model included the main effects of treatment, breed, age, cycling status, and their respective interactions. To compare all CIDR treatment groups against untreated ewes, contrast statements analyzed d to estrus, d to lambing, percent ewes exhibiting estrus, pregnancy rate, lambing rate and prolificacy to both the first service period and overall. For Experiments 1 and 2, repeated measures of the MIXED procedure of SAS were used to analyze serum concentrations of P4. Main effects of the model included treatment, time, cyclic status, and treatment × time interaction. Limited differences for concentrations of P4 within time were detected between CIDR treatments; therefore, results from all CIDR treatment groups were combined (CIDR-treated) and compared to untreated ewes. To better understand differences in P4 patterns between ewes that became pregnant during the first 17 d post ram introduction and those that did not, ewes were sorted into pregnant and non-pregnant groups. Data are presented as least squares means and treatment differences were considered significant at P ≤ 0.05. Interactions with a P-value > 0.20 were removed from the model.
Please cite this article in press as: Jackson, C.G., et al., Efficacy of various five-day estrous synchronization protocols in sheep. Small Ruminant Res. (2014), http://dx.doi.org/10.1016/j.smallrumres.2014.04.004
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3. Results 3.1. Experiment 1 3.1.1. Reproductive performance data Overall d to estrus after CIDR removal was greater (P ≤ 0.01; Table 1) in CON and CTP compared with CT, whereas CTG was intermediate. Surprisingly, no differences (P ≥ 0.11) were detected among treatments for d to lambing as well as for the proportion of ewes exhibiting estrus to both the first service period and overall. Consequently, no differences (P ≥ 0.27) were detected for the proportion of ewes becoming pregnant and lambing as well as prolificacy to both the first service period and overall among treatment groups. Contrasts comparing all ewes receiving a CIDR (CT, CTP, and CTG) with untreated ewes revealed that ewes receiving a CIDR exhibited estrus sooner (P = 0.01) after ram introduction than untreated ewes. Similarly, CIDR-treated ewes lambed earlier (P = 0.05) in the lambing season compared with untreated ewes. No differences (P ≥ 0.18) were detected for percent ewes exhibiting estrus, pregnancy rate, lambing rate and prolificacy to the overall breeding season for CIDR-treated ewes compared with untreated ewes. 3.1.2. Serum concentrations of progesterone in pregnant ewes Prior to treatment initiation, only 37% of ewes were determined to be fully cyclic. A treatment × day interaction (P = 0.01; Fig. 1) was present for concentrations of P4 in ewes that became pregnant (n = 89) during the first service period. Immediately, following CIDR removal (d 0), CIDR-treated ewes had greater (P ≤ 0.01) concentrations of P4 compared with untreated ewes. Untreated ewes had greater (P ≤ 0.01) concentrations of P4 on d 4 and 6 compared with CIDR-treated ewes. All CIDR-treated ewes had greater (P ≤ 0.02) concentrations of P4 compared with untreated ewes on d 11, 12, 14 and 20. 3.1.3. Serum concentrations of progesterone in non-pregnant ewes In non-pregnant ewes (n = 87), a treatment × day interaction (P = 0.01; Fig. 2) was detected. On d 0 and 8, CIDR-treated ewes had greater (P ≤ 0.01) concentrations of P4 compared with untreated ewes. Lastly, on d 17 and 20, untreated ewes had greater (P ≤ 0.01) concentrations of P4 compared with CIDR-treated ewes. 3.2. Experiment 2 3.2.1. Reproductive performance data Among treatments, CTG and CT treated ewes exhibited estrus in fewer days (P ≤ 0.02; Table 2) post-CIDR removal than CON ewes; whereas, CTP treated ewes did not differ (P = 0.07) from CON ewes. No differences (P = 0.23) were detected among treatment groups for days to lambing and for percentage of ewes exhibiting estrus during the first 17 d after ram introduction and overall. Pregnancy rate to the first 17 d post CIDR removal were similar (P = 0.79) for CON, CT, CTP, and CTG. No differences (P ≥ 0.47) were observed
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among treatments for overall pregnancy rate and lambing rate. Lastly, no differences (P ≥ 0.44) were detected for prolificacy to the first 17 d or overall among treatments. Contrasts comparing all ewes receiving a CIDR (CT, CTP, and CTG) with untreated ewes revealed that, CIDR-treated ewes exhibited estrus in less days after ram introduction (P = 0.04) than untreated ewes. However, no differences (P ≥ 0.33) were detected among CIDR-treated ewes and untreated ewes for days to lambing, the proportion of ewes exhibiting estrus, proportion of ewes pregnant, lambing rate, or prolificacy. 3.2.2. Serum concentrations of progesterone in pregnant ewes Prior to treatment initiation, a majority (62%) of ewes were classified as cyclic. A treatment × day interaction (P = 0.01; Fig. 3) for concentrations of P4 was detected in pregnant ewes (n = 32). Concentrations of P4 were greater (P ≤ 0.05) in untreated ewes compared with CIDR-treated ewes on d-12, -5, 2, and 3. On d 14 and 20, CIDR treated ewes had greater (P ≤ 0.02) concentrations of P4 compared with untreated ewes. 3.2.3. Serum concentrations of progesterone in non-pregnant ewes In non-pregnant ewes (n = 46), no interactions (P = 0.36, Fig. 4) were detected between CIDR-treated and untreated ewes. 4. Discussion Experiment 1: Due to the recent approval of CIDRs for use in the commercial sheep industry, with the intent of inducing estrus in ewes during the anestrous season, we set out to determine the efficacy of this new label recommendation during both the transition and anestrous seasons. Limited research has been conducted thus far utilizing the CIDR for the recommended insertion time. Therefore, due to the novelty of this study, comparisons were made against other research that utilized various exogenous P4 sources and insertion durations to induce estrus in order to allow us to determine the efficacy of the 5 d CIDR when used in a commercial setting. During the transition period (August through October) after the summer solstice, d to estrus were reduced by 6 d in CT ewes and by 3 d for all the remaining ewes receiving a 5 d CIDR treatment compared with CON ewes. This pattern of reduced interval to estrus is consistent with Wheaton et al. (1992) who reported a reduced interval to estrus in ewes treated with a CIDR (average 2 d) compared with untreated ewes (average 21 d). In this study, average d to lambing was reduced by 9 d for ewes treated with a 5 d CIDR (167 d) compared to CON ewes (176 d). Although Titi et al. (2010) reported no differences in d to lambing between those ewes treated with an exogenous P4 source for 5 d (average 168 d) and untreated ewes (174 d) the actual days to lambing for each treatment groups were similar. Although no differences were observed in the percentage of ewes exhibiting estrus overall, our findings are in close agreement with a study by Wheaton et al. (1992) in which 95% of untreated ewes and
Please cite this article in press as: Jackson, C.G., et al., Efficacy of various five-day estrous synchronization protocols in sheep. Small Ruminant Res. (2014), http://dx.doi.org/10.1016/j.smallrumres.2014.04.004
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Table 1 Reproductive performance of ewes after 5-d controlled internal drug releasing (CIDR) synchronization protocols administered during the transition season (Experiment 1). Treatment1
Variable
Number of ewes Days to estrus4 Days to lambing5
CIDR vs untreated P-value3 2
CON
CT
CTP
CTG
SEM
P-value
44 9.5a 176
45 3.6c 167
43 6.5b 167
44 6.2b,c 172
1.10 3.13
0.01 0.11
0.01 0.05
Ewes exhibiting estrus, %6 First service period7 Overall
67 86
89 96
77 96
75 89
6.60 4.30
0.28 0.43
0.17 0.25
Pregnancy rate, %8 First service period Overall
37 90
62 87
56 88
46 89
8.00 5.10
0.27 0.97
0.13 0.79
Lambing rate9 First service period Overall
0.68 1.52
0.87 1.27
0.90 1.42
0.65 1.26
0.14 0.12
0.55 0.41
0.50 0.21
Prolificacy10 First service period Overall
1.85 1.70
1.40 1.46
1.60 1.61
1.40 1.45
0.22 0.10
0.28 0.35
0.13 0.18
Values within the same row with different superscripts differ (P ≤ 0.05). 1 Treatments: CON = untreated; CT = CIDR inserted on d-5 and removed on d 0; CTP = 5 d CIDR and PG given at removal; CTG = GnRH given on d-5, 5 d CIDR and PG given at removal with ram exposure starting on d 0. 2 P-value for F-tests of the mean. 3 All CIDR based treatments (CT, CTP, and CTG) were compared to the untreated ewes (CON) with a contrast statement. 4 Days to estrus post ram introduction as indicated by marks from rams equipped with marking harnesses. 5 Days to lambing post CIDR removal. 6 Number of ewes marked by a ram as a percentage of all ewes treated. 7 First service period is defined as the first 17 d post CIDR removal. 8 Number of ewes that lambed less than 155 d post CIDR removal expressed as a percentage of all ewes treated. 9 Lambs born per ewe exposed. 10 Lambs born per ewe lambing. ab
100% of ewes receiving a CIDR for 12 d exhibited estrus overall. Pregnancy and lambing rates to the first service period and overall were not clearly improved by inclusion of a CIDR in any of the treatment protocols when compared with CON ewes in the current study. However, there were
mild indications that CIDR treatments increased pregnancy rates (55% vs 37%) and lowered prolificacy rates (1.85 vs 1.47) to the first service period compared to untreated ewes. However, CT ewes had an overall pregnancy rate of 87% and a lambing rate of 1.27 lambs born per ewe exposed. No comparable data was found utilizing ewes
trt*time Untreated CIDR-treated
9
0.05 †
8 †
7 Progesterone, ng/mL
P
† † †
6 5 4†
†
3 2 1 0
-12
-5
0 1 2 3 4 5 6 7 8 9 1011121314151617181920 Day
Fig. 1. Serum progesterone (P4) profiles before and after 5-d controlled internal drug release (CIDR) synchronization protocols administered during the transition period of pregnant ewes (n = 89) in Experiment 1. Treatments: untreated = untreated; all CIDR based treatments = CIDR-treated. Differences (P ≤ 0.05) between untreated and CIDR-treated ewes within day are denoted by †.
Please cite this article in press as: Jackson, C.G., et al., Efficacy of various five-day estrous synchronization protocols in sheep. Small Ruminant Res. (2014), http://dx.doi.org/10.1016/j.smallrumres.2014.04.004
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trt*time
Progesterone, ng/mL
Untreated CIDR-treated
5
P
0.05
6 5 †
4
†
†
3† 2 1 0 -5
-12
0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 Day
Fig. 2. Serum progesterone (P4) profiles before and after 5-d controlled internal drug release (CIDR) synchronization protocols administered during the transition period of non-pregnant ewes (n = 87) in Experiment 1. Treatments: untreated = untreated; all CIDR based treatments = CIDR-treated. Differences (P ≤ 0.05) between untreated and CIDR-treated ewes within day are denoted by †.
exhibiting estrous cyclicity; however, this contrasts with Knights et al. (2001b) who reported an overall increase in pregnancy rate by 41% and 0.26 lambs born per ewe exposed for lambing rate overall in ewes treated with a CIDR for 5-d compared to untreated ewes in the anestrous period. Conversely, Titi et al. (2010) reported an increase in prolificacy in ewes treated with GnRH and PG coupled
with an exogenous P4 source and we failed to detect differences in prolificacy to both the first service period and overall. Perhaps in the study conducted by Titi et al. (2010) when GnRH was administered, more ewes had less active corpora lutea, thus enhancing the follicular development and synchrony of those ewes. Further, ewes in the Titi et al. (2010) study may have previously experienced a
Table 2 Reproductive performance of ewes after 5-d controlled internal drug releasing (CIDR) synchronization protocols administered during the anestrous season (Experiment 2). Variable
Treatment1
CIDR vs Untreated P-value3
CON
CT
CTP
CTG
Number of ewes Days to estrus4 Days to lambing5
16 6.5a 152
21 3.4bc 152
20 5.9ac 154
21 3.0b 150
Ewes exhibiting estrus, %6 First service period7 Overall
94 100
78 82
90 100
96 96
42 42
37 39
35 55
50 58
Pregnancy rate, %8 First service period Overall
SEM
P-value2
1.05 2.99
0.02 0.55
0.04 0.95
8.00 6.00
0.41 0.23
0.54 0.33
0.79 0.59
0.91 0.56
12.4 12.1
Lambing rate9 First service period Overall
0.48 0.47
0.50 0.51
0.55 0.85
0.63 0.71
0.17 0.18
0.90 0.47
0.72 0.36
Prolificacy10 First service period Overall
1.17 1.15
1.40 1.31
1.38 1.55
1.20 1.24
0.19 0.19
0.69 0.44
0.49 0.36
Values within the same row with different superscripts differ (P ≤ 0.05). 1 Treatments: CON = untreated; CT = CIDR inserted on d-5 and removed on d 0; CTP = 5 d CIDR and PG given at removal; CTG = GnRH given on d-5, 5 d CIDR and PG given at removal with ram exposure starting on d 0. 2 P-value for F-tests of the mean. 3 All CIDR based treatments (CT, CTP, and CTG) were compared to the untreated ewes (CON) with a contrast statement. 4 Days to estrus post ram introduction as indicated by marks from rams equipped with marking harnesses. 5 Days to lambing post CIDR removal. 6 Number of ewes marked by a ram as a percentage of all ewes treated. 7 First service period is defined as the first 17 d post CIDR removal. 8 Number of ewes that lambed less than 155 d post CIDR removal expressed as a percentage of all ewes treated. 9 Lambs born per ewe exposed. 10 Lambs born per ewe lambing. ab
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Progesterone, ng/mL
Untreated CIDR-treated
trt*time
11
†
9 7
†
0.05
10 8
†
P
† †
†
6 5 4 3 2 1 0
-12
0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 Day
-5
Fig. 3. Serum progesterone (P4) profiles before and after 5-d controlled internal drug release (CIDR) synchronization protocols administered during the anestrous season of pregnant ewes (n = 32) in Experiment 2. Treatments: untreated = untreated; all CIDR based treatments = CIDR-treated. Differences (P ≤ 0.05) between untreated and CIDR-treated ewes within day are denoted by †.
false heat or actually expressed a full estrus cycle prior to the administration of PG. In our experiment, we theorize that prolificacy rates were higher in the control ewes because they did not conceive to the first estrous cycle. Ewe lambing rate (prolificacy) is greater in ewes that lamb later in the lambing season (Glimp, 1971). Through the use of exogenous P4, the lifespan of the corpus luteum may be manipulated to aid in regulating estrus and ovulation (Hansel and Convey, 1983). The CIDR provided exogenous P4 as evident by elevated P4 concentrations on d 0 in CIDR-treated ewes regardless of their pregnancy status. In contrast, ewes not receiving a CIDR had greater P4 concentrations from d 4 to 6 in pregnant ewes. This decrease in concentrations of P4 from d 4 to 6 for CIDR-treated ewes likely indicates that a greater proportion of CIDR-treated ewes were either in anestrous, proestrus, or estrus compared with ewes not receiving a CIDR. The reduced concentrations of P4 compliment the difference found in that CIDR-treated had reduced d to estrus compared with ewes not receiving a CIDR. Similar to Bartlewski et al. (1999) who reported concentrations of P4 begin to increase between 3 and 7 7
CIDR-treated
6
Progesterone, ng/mL
Untreatd
d after estrus and then reach peak concentrations at approximately d 12, we found in the current study that CIDR-treated ewes when compared with untreated ewes follow this fluctuation of increased concentrations of P4 rising after d 6. However, peak levels of concentrations of P4 are seen at approximately d 20 for those ewes who were determined to be pregnant. Seasonal, lactational, and prepubertal ewes that are isolated from rams for at least a month, will exhibit a synchronized heat after re-introduction to the male approximately one to one and a half estrous cycles (17–24 d) later (Underwood et al., 1944; Martin et al., 1986). This has been termed the ‘ram effect’ (Underwood et al., 1944; Martin et al., 1986). It appears untreated ewes that become pregnant in the first service period were likely influenced by the ram effect which is evident by a delayed increase in concentrations of P4 compared with CIDR-treated ewes. This interval was relatively similar to that reported by Schinckel (1954), who indicated an interval of 14 d after ram introduction. Experiment 2: During the anestrous period (April through May) in which this study was conducted, CT and
trt*time
P = 0.36
5 4 3 2 1 0
-12
-5
0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 Day
Fig. 4. Serum progesterone (P4) profiles before and after 5-d controlled internal drug release (CIDR) synchronization protocols administered during the anestrous season of non-pregnant ewes (n = 46) in Experiment 2. Treatments: untreated = untreated; all CIDR based treatments = CIDR-treated.
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CTG ewes came into estrus more quickly after ram introduction than the CTP and CON ewes. This data indicates that CT and CTG treated ewes displayed a tighter synchrony to estrus. Similarly, Wheaton et al. (1992) observed ewes treated with a CIDR having a reduced interval to estrus compared with untreated ewes. In the current study, no differences were detected in overall days to lambing among treatment groups as well as all CIDR-treated ewes compared with CON. Conversely, Knights et al. (2001a) reported 42% of anestrous ewes treated with a 5 d CIDR lambed approximately 12–13 d earlier than untreated ewes. Similar to Knights et al. (2001a) and Wheaton et al. (1993) reported ewes in anestrous who were treated with a CIDR for 12 d required fewer d (157 ± 2 d) to lamb after introduction of rams compared with control ewes (170 ± 2 d). No statistical differences were detected among treatments for percentage of ewes exhibiting estrus to the first service and overall in the current study which is consistent with Wheaton et al. (1992) who reported no differences in ewes overall exhibiting estrus between untreated ewes and those receiving a CIDR insert for 12 d. Although we attempted to get all ewes in the anestrous period, prior to imposing treatments, ewes in the untreated group were already cycling as evidence by progesterone concentrations ≥1.0 ng/mL. Therefore, it is not surprising that pregnancy rates within the first service period after CIDR removal were similar in the current study for all treatments. In contrast, Knights et al. (2001a,b) documented greater pregnancy rates and an increase in lambing rate (Knights et al., 2001a) in P4 treated ewes compared with control ewes. In contrast to the current study, Titi et al. (2010) reported an increase in prolificacy in ewes treated with exogenous P4 when coupled with the use of GnRH and PG. Although no differences were detected for overall reproductive performance data, this study suggests that the CTG treatment was the most effective at concentrating estrus shortly after CIDR removal compared with CON, CT, and CTP treatments. In the present study, CIDR-treated ewes that were determined pregnant to the first service period had reduced concentrations of P4 from d 0 to 5. Perhaps this reduction in concentrations of P4 indicates decreased luteal activity which led to a more uniform synchronization to estrus further into the estrous cycle. Cyclicity during the anestrous season is not uncommon for the Dorset and Katahdin breeds as indicated by concentrations of P4 ≥ 1.0 ng/mL in 62% of the CON ewes; however, the overall pregnancy rate of 42% for CON ewes indicates CON ewes were not fully cyclic. It has been documented that a semi-cyclic pattern in ovarian activity is evident during early and late anestrous ewes (Cole and Miller, 1935). In the non-pregnant ewes, ewes had concentrations of P4 below 2.0 ng/mL from d 2 to 8. Both untreated and CIDR-treated ewes may have been in such a deep anestrous state that the treatments and/or ram effect were not powerful enough to bring them out of anestrous. Perhaps it did for a short period, explaining why the CIDR-treated ewes experienced an increase in concentrations of P4 around d 10; however, reverting back to decreased concentrations of P4 by d 15. Likewise, with the untreated ewes, an increase and then steady decline in concentrations of P4 can be observed from d
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1 to 14, with concentrations of P4 increasing again at d 15. Perhaps ram introduction stimulated cyclicity in the untreated ewes. These results are consistent with Martin et al. (1986), who reported that after 6 d of the short cycle, a second ovulation is induced followed by a fertile cycle of normal length. This apparent effect of ram introduction provides justification that a majority of the ewes were either non-cyclic or semi-cyclic. 5. Conclusions In the current study, during the transition and anestrous season 5 d CIDR inserts decreased overall days to detected estrus. Ewe CIDR-treatments decreased days to lambing in ewes bred during August and September. Regardless of season, inclusion of GnRH and PG into 5 d CIDR protocols did not improve ewe reproductive performance. The 5 d CIDR, regardless of protocol, did not markedly influence measures of reproductive performance desired by commercial sheep producers. The efficacy of CIDR protocols to synchronize estrus and improve fertility during the transition or anestrous period was not clearly demonstrated in this study. Conflict of interest None declared. Acknowledgements This study was supported by Zoetis Animal Health and the North Dakota State Board of Agriculture Research and Education Grant # 11-28. We thank the NDSU Sheep Shepherd Skip Anderson for all of his assistance during these studies. References Bartlewski, P.M., Beard, A.P., Rawlings, N.C., 1999. An ultrasonographic study of luteal function in breeds of sheep with different ovulation rates. Theriogenology 52, 115–130. Carlson, K.M., Pohl, H.A., Marcek, J.M., Muser, R.K., Wheaton, J.E., 1989. Evaluation of progesterone controlled internal drug release dispensers for synchronization of estrus in sheep. Anim. Reprod. Sci. 18, 205–218. Cole, H.H., Miller, R.F., 1935. Changes in the reproductive organs of the ewe with some data bearing on their control. Am. J. Anat. 57, 39–97. Glimp, H.A., 1971. Effect of breed and mating season on reproductive performance of sheep. J. Anim. Sci. 32, 1176–1182. Hamra, A.H., Massri, Y.G., Marcek, J.M., Wheaton, J.E., 1986. Plasma progesterone levels in ewes treated with progesterone-controlled internal drug-release dispensers, implants and sponges. Anim. Reprod. Sci. 11, 187–194. Hansel, W., Convey, E.M., 1983. Physiology of the estrous cycle. J. Anim. Sci. 57, 404–424. Knights, M., Hoehn, T., Lewis, P.E., Inskeep, E.K., 2001a. Effectiveness of intravaginal progesterone inserts and FSH for inducing synchronized estrus and increasing lambing rate in anestrous ewes. J. Anim. Sci. 79, 1120–1131. Knights, M., Maze, T.D., Bridges, P.J., Lewis, P.E., Inskeep, E.K., 2001b. Shortterm treatment with a controlled internal drug releasing (CIDR) device and FSH to induce fertile estrus and increase prolificacy in anestrous ewes. Theriogenology 55, 1181–1191. Martin, G.B., Oldham, C.M., Cognie, Y., Pearce, D.T., 1986. The physiological responses of anovulatory ewes to the introduction of rams – a review. Livest. Prod. Sci. 15, 219–247. Schinckel, P.G., 1954. The effect of the presence of the ram on the ovarian activity of the ewe. Aust. J. Agric. Res. 5, 465–469.
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Schneider, F.A., Hallford, D.M., 1996. Use of a rapid progesterone radioimmunoassay to predict pregnancy and fetal numbers in ewes. Sheep Goat Res. J. 12, 33–38. Titi, H.H., Kridli, R.T., Alnimer, M.A., 2010. Estrus synchronization in sheep and goats using combinations of GnRH, progestagen and prostaglandin F2␣ . Reprod. Domest. Anim. 45, 594–599. Underwood, E.J., Shier, F.L., Davenport, N., 1944. Studies in sheep husbandry in W.A. V. The breeding season of Merino, Crossbred and British breed ewes in the agricultural districts. J. Agric. West. Aust. 2, 135–143.
Wheaton, J.E., Windels, H.F., Johnston, L.J., 1992. Accelerated lambing using exogenous progesterone and the ram effect. J. Anim. Sci. 70, 2628–2635. Wheaton, J.E., Carlson, K.M., Windels, H.F., Johnston, L.J., 1993. CIDR: a new progesterone-releasing intravaginal device for induction of estrus and cycle control in sheep and goats. Anim. Reprod. Sci. 33, 127–141. Wildeus, S., 2000. Current concepts in synchronization of estrus: sheep and goats. J. Anim. Sci. 77, 1–14.
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Contents lists available at ScienceDirect
Small Ruminant Research journal homepage: www.elsevier.com/locate/smallrumres
Efficacy of various five-day estrous synchronization protocols in sheep C.G. Jackson a , T.L. Neville a , V.R.G. Mercadante b , K.M. Waters b , G.C. Lamb b , C.R. Dahlen a , R.R. Redden a,∗ a b
Department of Animal Sciences, North Dakota State University, NDSU Dept. 7630, P.O. Box 6050, Fargo, ND 58108-6050, USA North Florida Research and Education Center, University of Florida, 3925 HWY 71, Marianna, FL 32446-8091, USA
a r t i c l e
i n f o
Article history: Received 26 February 2014 Accepted 8 April 2014 Available online xxx
Keywords: Controlled internal drug releasing inserts Estrous synchronization Sheep
a b s t r a c t The objective of this study was to determine the effects of ewe estrus synchronization protocols including combinations of controlled internal drug releasing (CIDR) inserts for 5 d, GnRH, and PG on concentrations of progesterone (P4) and reproductive performance of ewes during different breeding seasons. Experiment 1 was conducted during the transition season (August–October) and Experiment 2 was conducted during the anestrous season (April–May). In both experiments ewes received 1 of 4 treatments: (1) untreated (CON); (2) CIDR (0.3 g P4) insert (CT); (3) CIDR and PG at CIDR removal (CTP); or (4) GnRH at CIDR insertion and PG at CIDR removal (CTG). Rams equipped with marking harnesses were introduced at CIDR removal (d 0) and ewes were observed for breeding marks twice daily at 0800 h and 1700 h. In Experiment 1, days to estrus was greater (P ≤ 0.01) in CON (9.5 ± 1.10 d) and CTP (6.5 ± 1.04 d) compared with CT (3.6 ± 1.03 d). No differences (P = 0.27) were detected in pregnancy rate within the first 17 d of the breeding season for CON, CT, CTP, and CTG treatments (37 ± 7.8%, 62 ± 7.5%, 56 ± 8.1%, and 46 ± 7.7%, respectively). Similarly, no differences (P ≥ 0.28) were detected among treatments for lambing rate and prolificacy. Contrasts for all CIDR treatments compared to controls determined that CIDR treated ewes had less days to estrus and lambing (P ≤ 0.05) than control ewes. In Experiment 2, days to estrus was greater (P ≤ 0.02) in CON (6.5 ± 1.05 d) and CTP (5.9 ± 0.88 d) compared with CTG (3.0 ± 0.88 d). No differences were detected (P ≥ 0.23) among treatments for overall percentage of ewes exhibiting estrus. No differences (P = 0.79) in pregnancy rate within the first 17 d of the breeding season were detected among treatments (42 ± 12.4%, 37 ± 11.3%, 35 ± 11.1%, and 50 ± 10.9% for CON, CT, CTP, and CTG, respectively). No differences (P ≥ 0.44) were observed among treatments in overall lambing rate and prolificacy. In conclusion, inserting a CIDR for 5 d prior to ram introduction regardless of the use of PG or GnRH, reduced the interval to estrus compared with untreated ewes but did not impact overall pregnancy rates, lambing rates, or prolificacy. Published by Elsevier B.V.
1. Introduction
∗ Corresponding author. Tel.: +1 701 231 5597; fax: +1 701 231 7590. E-mail address: [email protected] (R.R. Redden).
Reproductive performance is one of the largest determining factors of profitability in today’s sheep industry and will aid in its future success and long-term sustainability. For the U.S. sheep industry to remain competitive in
http://dx.doi.org/10.1016/j.smallrumres.2014.04.004 0921-4488/Published by Elsevier B.V.
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the market place and to increase year-round demand for lamb, producers could benefit from technology that would effectively synchronize estrus in ewes during the transition and anestrous seasons. Controlled internal drug releasing (CIDR) inserts were approved for use in sheep by the U.S. Food and Drug Administration in 2009 for estrous synchronization and are currently labeled for 5 d use to induce estrus in anestrous ewes. Due to the seasonal nature of estrous cyclicity of most sheep, exogenous hormones can be used to facilitate the induction and synchronization of estrus in the anestrous period (Carlson et al., 1989). This is of particular interest to those who produce lambs consistently throughout the year in an accelerated lambing system. Producers in a traditional spring lambing system can also benefit from exogenous progesterone (P4) and PG use to synchronize estrus during the transition period (when the amount of day light begins to decrease in the northern and southern latitudes and ewes begin to exhibit estrous cyclicity). Due to the seasonal nature of ovine estrous cyclicity, PG is only effective in animals already cycling; therefore, only a portion of ewes may respond to this treatment. To assist in initiating hormonal events necessary to synchronize estrous cycles, such as estrus and ovulation, GnRH may also be incorporated, especially in anestrous ewes (Carlson et al., 1989; Wildeus, 2000). Therefore, our objective was to determine the efficacy of CIDR insertion for 5 d when used alone or in combination with GnRH and PG during both the transition and anestrous seasons. We hypothesized that estrous synchronization with CIDR, GnRH, and/or PG will decrease days to estrus and increase pregnancy, lambing, and prolificacy rates. 2. Materials and methods The North Dakota State University Institutional Animal Care and Use Committee approved all protocols and procedures involving animals used in this research under protocol A11007. Animals were housed at the North Dakota State University Sheep Unit in Fargo, ND. 2.1. Animals and treatments Experiment 1: In August 2010 and 2011, Columbia (n = 38 and 38, respectively) and Hampshire (n = 47 and 53, respectively) ewes were randomly assigned to 1 of 4 treatments: (1) untreated (CON; n = 44); (2) 5 d CIDR insert (EAZI-BREED CIDR Sheep Insert, 0.3 g P4, Zoetis Animal Health, Florham Park, NJ; CT; n = 45); (3) 5 d CIDR and PG (Lutalyse, 10 mg dinoprost tromethamine i.m., Zoetis Animal Health) at CIDR removal (CTP; n = 44); and (4) GnRH (Factrel, 0.025 mg gonadorelin hydrochloride i.m., Zoetis Animal Health) at CIDR insertion and PG at CIDR removal (CTG; n = 43). Treatments were assigned over 3 time periods, 1 wk apart, to ensure an adequate ram to ewe ratio throughout the breeding period. One week prior to treatment application, ewes were moved to paddocks that had not been previously grazed for 1 mo. Upon treatment administration, all ewes were moved to a drylot with continuous access to fresh water and received alfalfa hay (3 kg/ewe) and a 14% CP concentrate ration (1 kg/ewe) daily. Prior to treatment initiation, ewes were isolated from males. Rams equipped with marking harnesses were single sire mated to ewes within treatment group at a ewe to ram ratio ≤25:1. Ewes were exposed to rams at CIDR insert removal (d 0) for a 60-d breeding season. Ewes were observed twice daily for breeding marks at 0800 h and 1700 h starting on d 0. Ewes were determined to be in estrus as indicated by breeding marks (when ≥60% of the rump was colored with wax marker). Experiment 2: In April 2011, during the anestrous period, multiparous and nulliparous Dorset and Katahdin (n = 61 and n = 17, respectively) ewes from the North Dakota State University Sheep Unit in Fargo, ND, were
randomly assigned to 1 of 4 treatments as described previously in Experiment 1: (1) untreated (CON; n = 16); (2) CIDR inserted for 5 d (CT; n = 21); (3) CIDR inserted for 5 d and PG at removal (CTP; n = 20); and (4) GnRH given at CIDR insertion and PG at CIDR removal (CTG; n = 21). Prior to treatment application, ewes were managed as a common group in a dry lot, isolated from all males, and had ad libitum access to alfalfa hay and fresh water. On d 0, 4 rams equipped with marking harnesses were introduced to ewes immediately post CIDR removal with a ewe to ram ratio ≤25:1. Beginning at d 0, ewes were observed for breeding marks twice daily at 0800 h and 1700 h and were determined to be in estrus as indicated by breeding marks. The breeding season lasted for duration of 42 d. 2.2. Reproductive performance data collection For both experiments data collected included days to estrus, days to lambing, percentage of ewes exhibiting estrus (divided the number of ewes marked by a ram by the total number of ewes on study), pregnancy rate (number of ewes lambing as a percentage of ewes in all treatments), lambing rate (number of lambs/ewe exposed), and prolificacy (number of lambs/ewe lambing). The latter 4 measures of interest were further broken down to represent the length of an estrous cycle (first 17 d of the breeding season), or the entire breeding season (overall; from d 0 until ram removal). 2.3. Blood collection and analysis To evaluate concentrations of serum P4 for Experiments 1 and 2, blood samples were collected via jugular venipuncture into 10-mL serum tubes (BD Vacutainer Serum, Becton, Dickinson and Company, Franklin Lakes, NJ) and immediately placed on ice. Samples were centrifuged at 4 ◦ C for 30 min at 1500 × g. Then serum was transferred into plastic 2.0-mL microcentrifuge tubes and frozen at −20 ◦ C until assayed. In Experiment 1, blood samples were collected 1 wk prior to (d-12) and on the day of treatment initiation (d-5) to obtain baseline concentrations of P4. Additional samples were collected at CIDR removal (d 0) and on d 2, 4, 6, 8, 10, and 12 in 2010 and on d 1, 2, 3, 4, 5, 8, 11, 14, 17, and 20 in 2011. In Experiment 2, samples were collected on d-12, -5, 0, 1, 2, 3, 4, 5, 8, 11, 14, 17, and 20 relative to CIDR removal. To detect circulating concentrations of P4, samples were assayed using a solid-phase, no extraction RIA (Coat-a-Count Progesterone, Diagnostic Products Corporation, Los Angeles, CA). The assay kit was validated for ovine serum using an assay volume of 100 L (Hamra et al., 1986; Schneider and Hallford, 1996). Assay was standardized to a curve of 0.1, 0.25, 0.5, 1.0, 2.0, 5.0, 10.0, and 20.0 ng/mL. Assay sensitivity for a 100-L sample was 0.1 ng/mL. Ewes were determined to be cycling if serum P4 concentrations were greater than 1 ng/mL on either or both the d-12 or -5 samples. All samples for a single ewe were analyzed within the same assay and treatments were run in a random order. The intraand inter-assay CVs for Experiment 1 were 5.39% and 3.61%, respectively and were 2.28% and 2.21%, respectively for Experiment 2. 2.4. Statistical analysis The MIXED procedures of SAS 9.2 (SAS Inst., Inc., Cary, NC) were utilized to examine d to detected estrus and d to lambing. The model included the main effects of treatment, breed, age, cycling status, and their respective interactions. The GLIMMIX procedures of SAS were used to analyze percentage of ewes exhibiting estrus, pregnancy rate, lambing rate, and prolificacy. The model included the main effects of treatment, breed, age, cycling status, and their respective interactions. To compare all CIDR treatment groups against untreated ewes, contrast statements analyzed d to estrus, d to lambing, percent ewes exhibiting estrus, pregnancy rate, lambing rate and prolificacy to both the first service period and overall. For Experiments 1 and 2, repeated measures of the MIXED procedure of SAS were used to analyze serum concentrations of P4. Main effects of the model included treatment, time, cyclic status, and treatment × time interaction. Limited differences for concentrations of P4 within time were detected between CIDR treatments; therefore, results from all CIDR treatment groups were combined (CIDR-treated) and compared to untreated ewes. To better understand differences in P4 patterns between ewes that became pregnant during the first 17 d post ram introduction and those that did not, ewes were sorted into pregnant and non-pregnant groups. Data are presented as least squares means and treatment differences were considered significant at P ≤ 0.05. Interactions with a P-value > 0.20 were removed from the model.
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3. Results 3.1. Experiment 1 3.1.1. Reproductive performance data Overall d to estrus after CIDR removal was greater (P ≤ 0.01; Table 1) in CON and CTP compared with CT, whereas CTG was intermediate. Surprisingly, no differences (P ≥ 0.11) were detected among treatments for d to lambing as well as for the proportion of ewes exhibiting estrus to both the first service period and overall. Consequently, no differences (P ≥ 0.27) were detected for the proportion of ewes becoming pregnant and lambing as well as prolificacy to both the first service period and overall among treatment groups. Contrasts comparing all ewes receiving a CIDR (CT, CTP, and CTG) with untreated ewes revealed that ewes receiving a CIDR exhibited estrus sooner (P = 0.01) after ram introduction than untreated ewes. Similarly, CIDR-treated ewes lambed earlier (P = 0.05) in the lambing season compared with untreated ewes. No differences (P ≥ 0.18) were detected for percent ewes exhibiting estrus, pregnancy rate, lambing rate and prolificacy to the overall breeding season for CIDR-treated ewes compared with untreated ewes. 3.1.2. Serum concentrations of progesterone in pregnant ewes Prior to treatment initiation, only 37% of ewes were determined to be fully cyclic. A treatment × day interaction (P = 0.01; Fig. 1) was present for concentrations of P4 in ewes that became pregnant (n = 89) during the first service period. Immediately, following CIDR removal (d 0), CIDR-treated ewes had greater (P ≤ 0.01) concentrations of P4 compared with untreated ewes. Untreated ewes had greater (P ≤ 0.01) concentrations of P4 on d 4 and 6 compared with CIDR-treated ewes. All CIDR-treated ewes had greater (P ≤ 0.02) concentrations of P4 compared with untreated ewes on d 11, 12, 14 and 20. 3.1.3. Serum concentrations of progesterone in non-pregnant ewes In non-pregnant ewes (n = 87), a treatment × day interaction (P = 0.01; Fig. 2) was detected. On d 0 and 8, CIDR-treated ewes had greater (P ≤ 0.01) concentrations of P4 compared with untreated ewes. Lastly, on d 17 and 20, untreated ewes had greater (P ≤ 0.01) concentrations of P4 compared with CIDR-treated ewes. 3.2. Experiment 2 3.2.1. Reproductive performance data Among treatments, CTG and CT treated ewes exhibited estrus in fewer days (P ≤ 0.02; Table 2) post-CIDR removal than CON ewes; whereas, CTP treated ewes did not differ (P = 0.07) from CON ewes. No differences (P = 0.23) were detected among treatment groups for days to lambing and for percentage of ewes exhibiting estrus during the first 17 d after ram introduction and overall. Pregnancy rate to the first 17 d post CIDR removal were similar (P = 0.79) for CON, CT, CTP, and CTG. No differences (P ≥ 0.47) were observed
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among treatments for overall pregnancy rate and lambing rate. Lastly, no differences (P ≥ 0.44) were detected for prolificacy to the first 17 d or overall among treatments. Contrasts comparing all ewes receiving a CIDR (CT, CTP, and CTG) with untreated ewes revealed that, CIDR-treated ewes exhibited estrus in less days after ram introduction (P = 0.04) than untreated ewes. However, no differences (P ≥ 0.33) were detected among CIDR-treated ewes and untreated ewes for days to lambing, the proportion of ewes exhibiting estrus, proportion of ewes pregnant, lambing rate, or prolificacy. 3.2.2. Serum concentrations of progesterone in pregnant ewes Prior to treatment initiation, a majority (62%) of ewes were classified as cyclic. A treatment × day interaction (P = 0.01; Fig. 3) for concentrations of P4 was detected in pregnant ewes (n = 32). Concentrations of P4 were greater (P ≤ 0.05) in untreated ewes compared with CIDR-treated ewes on d-12, -5, 2, and 3. On d 14 and 20, CIDR treated ewes had greater (P ≤ 0.02) concentrations of P4 compared with untreated ewes. 3.2.3. Serum concentrations of progesterone in non-pregnant ewes In non-pregnant ewes (n = 46), no interactions (P = 0.36, Fig. 4) were detected between CIDR-treated and untreated ewes. 4. Discussion Experiment 1: Due to the recent approval of CIDRs for use in the commercial sheep industry, with the intent of inducing estrus in ewes during the anestrous season, we set out to determine the efficacy of this new label recommendation during both the transition and anestrous seasons. Limited research has been conducted thus far utilizing the CIDR for the recommended insertion time. Therefore, due to the novelty of this study, comparisons were made against other research that utilized various exogenous P4 sources and insertion durations to induce estrus in order to allow us to determine the efficacy of the 5 d CIDR when used in a commercial setting. During the transition period (August through October) after the summer solstice, d to estrus were reduced by 6 d in CT ewes and by 3 d for all the remaining ewes receiving a 5 d CIDR treatment compared with CON ewes. This pattern of reduced interval to estrus is consistent with Wheaton et al. (1992) who reported a reduced interval to estrus in ewes treated with a CIDR (average 2 d) compared with untreated ewes (average 21 d). In this study, average d to lambing was reduced by 9 d for ewes treated with a 5 d CIDR (167 d) compared to CON ewes (176 d). Although Titi et al. (2010) reported no differences in d to lambing between those ewes treated with an exogenous P4 source for 5 d (average 168 d) and untreated ewes (174 d) the actual days to lambing for each treatment groups were similar. Although no differences were observed in the percentage of ewes exhibiting estrus overall, our findings are in close agreement with a study by Wheaton et al. (1992) in which 95% of untreated ewes and
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Table 1 Reproductive performance of ewes after 5-d controlled internal drug releasing (CIDR) synchronization protocols administered during the transition season (Experiment 1). Treatment1
Variable
Number of ewes Days to estrus4 Days to lambing5
CIDR vs untreated P-value3 2
CON
CT
CTP
CTG
SEM
P-value
44 9.5a 176
45 3.6c 167
43 6.5b 167
44 6.2b,c 172
1.10 3.13
0.01 0.11
0.01 0.05
Ewes exhibiting estrus, %6 First service period7 Overall
67 86
89 96
77 96
75 89
6.60 4.30
0.28 0.43
0.17 0.25
Pregnancy rate, %8 First service period Overall
37 90
62 87
56 88
46 89
8.00 5.10
0.27 0.97
0.13 0.79
Lambing rate9 First service period Overall
0.68 1.52
0.87 1.27
0.90 1.42
0.65 1.26
0.14 0.12
0.55 0.41
0.50 0.21
Prolificacy10 First service period Overall
1.85 1.70
1.40 1.46
1.60 1.61
1.40 1.45
0.22 0.10
0.28 0.35
0.13 0.18
Values within the same row with different superscripts differ (P ≤ 0.05). 1 Treatments: CON = untreated; CT = CIDR inserted on d-5 and removed on d 0; CTP = 5 d CIDR and PG given at removal; CTG = GnRH given on d-5, 5 d CIDR and PG given at removal with ram exposure starting on d 0. 2 P-value for F-tests of the mean. 3 All CIDR based treatments (CT, CTP, and CTG) were compared to the untreated ewes (CON) with a contrast statement. 4 Days to estrus post ram introduction as indicated by marks from rams equipped with marking harnesses. 5 Days to lambing post CIDR removal. 6 Number of ewes marked by a ram as a percentage of all ewes treated. 7 First service period is defined as the first 17 d post CIDR removal. 8 Number of ewes that lambed less than 155 d post CIDR removal expressed as a percentage of all ewes treated. 9 Lambs born per ewe exposed. 10 Lambs born per ewe lambing. ab
100% of ewes receiving a CIDR for 12 d exhibited estrus overall. Pregnancy and lambing rates to the first service period and overall were not clearly improved by inclusion of a CIDR in any of the treatment protocols when compared with CON ewes in the current study. However, there were
mild indications that CIDR treatments increased pregnancy rates (55% vs 37%) and lowered prolificacy rates (1.85 vs 1.47) to the first service period compared to untreated ewes. However, CT ewes had an overall pregnancy rate of 87% and a lambing rate of 1.27 lambs born per ewe exposed. No comparable data was found utilizing ewes
trt*time Untreated CIDR-treated
9
0.05 †
8 †
7 Progesterone, ng/mL
P
† † †
6 5 4†
†
3 2 1 0
-12
-5
0 1 2 3 4 5 6 7 8 9 1011121314151617181920 Day
Fig. 1. Serum progesterone (P4) profiles before and after 5-d controlled internal drug release (CIDR) synchronization protocols administered during the transition period of pregnant ewes (n = 89) in Experiment 1. Treatments: untreated = untreated; all CIDR based treatments = CIDR-treated. Differences (P ≤ 0.05) between untreated and CIDR-treated ewes within day are denoted by †.
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trt*time
Progesterone, ng/mL
Untreated CIDR-treated
5
P
0.05
6 5 †
4
†
†
3† 2 1 0 -5
-12
0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 Day
Fig. 2. Serum progesterone (P4) profiles before and after 5-d controlled internal drug release (CIDR) synchronization protocols administered during the transition period of non-pregnant ewes (n = 87) in Experiment 1. Treatments: untreated = untreated; all CIDR based treatments = CIDR-treated. Differences (P ≤ 0.05) between untreated and CIDR-treated ewes within day are denoted by †.
exhibiting estrous cyclicity; however, this contrasts with Knights et al. (2001b) who reported an overall increase in pregnancy rate by 41% and 0.26 lambs born per ewe exposed for lambing rate overall in ewes treated with a CIDR for 5-d compared to untreated ewes in the anestrous period. Conversely, Titi et al. (2010) reported an increase in prolificacy in ewes treated with GnRH and PG coupled
with an exogenous P4 source and we failed to detect differences in prolificacy to both the first service period and overall. Perhaps in the study conducted by Titi et al. (2010) when GnRH was administered, more ewes had less active corpora lutea, thus enhancing the follicular development and synchrony of those ewes. Further, ewes in the Titi et al. (2010) study may have previously experienced a
Table 2 Reproductive performance of ewes after 5-d controlled internal drug releasing (CIDR) synchronization protocols administered during the anestrous season (Experiment 2). Variable
Treatment1
CIDR vs Untreated P-value3
CON
CT
CTP
CTG
Number of ewes Days to estrus4 Days to lambing5
16 6.5a 152
21 3.4bc 152
20 5.9ac 154
21 3.0b 150
Ewes exhibiting estrus, %6 First service period7 Overall
94 100
78 82
90 100
96 96
42 42
37 39
35 55
50 58
Pregnancy rate, %8 First service period Overall
SEM
P-value2
1.05 2.99
0.02 0.55
0.04 0.95
8.00 6.00
0.41 0.23
0.54 0.33
0.79 0.59
0.91 0.56
12.4 12.1
Lambing rate9 First service period Overall
0.48 0.47
0.50 0.51
0.55 0.85
0.63 0.71
0.17 0.18
0.90 0.47
0.72 0.36
Prolificacy10 First service period Overall
1.17 1.15
1.40 1.31
1.38 1.55
1.20 1.24
0.19 0.19
0.69 0.44
0.49 0.36
Values within the same row with different superscripts differ (P ≤ 0.05). 1 Treatments: CON = untreated; CT = CIDR inserted on d-5 and removed on d 0; CTP = 5 d CIDR and PG given at removal; CTG = GnRH given on d-5, 5 d CIDR and PG given at removal with ram exposure starting on d 0. 2 P-value for F-tests of the mean. 3 All CIDR based treatments (CT, CTP, and CTG) were compared to the untreated ewes (CON) with a contrast statement. 4 Days to estrus post ram introduction as indicated by marks from rams equipped with marking harnesses. 5 Days to lambing post CIDR removal. 6 Number of ewes marked by a ram as a percentage of all ewes treated. 7 First service period is defined as the first 17 d post CIDR removal. 8 Number of ewes that lambed less than 155 d post CIDR removal expressed as a percentage of all ewes treated. 9 Lambs born per ewe exposed. 10 Lambs born per ewe lambing. ab
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6
Progesterone, ng/mL
Untreated CIDR-treated
trt*time
11
†
9 7
†
0.05
10 8
†
P
† †
†
6 5 4 3 2 1 0
-12
0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 Day
-5
Fig. 3. Serum progesterone (P4) profiles before and after 5-d controlled internal drug release (CIDR) synchronization protocols administered during the anestrous season of pregnant ewes (n = 32) in Experiment 2. Treatments: untreated = untreated; all CIDR based treatments = CIDR-treated. Differences (P ≤ 0.05) between untreated and CIDR-treated ewes within day are denoted by †.
false heat or actually expressed a full estrus cycle prior to the administration of PG. In our experiment, we theorize that prolificacy rates were higher in the control ewes because they did not conceive to the first estrous cycle. Ewe lambing rate (prolificacy) is greater in ewes that lamb later in the lambing season (Glimp, 1971). Through the use of exogenous P4, the lifespan of the corpus luteum may be manipulated to aid in regulating estrus and ovulation (Hansel and Convey, 1983). The CIDR provided exogenous P4 as evident by elevated P4 concentrations on d 0 in CIDR-treated ewes regardless of their pregnancy status. In contrast, ewes not receiving a CIDR had greater P4 concentrations from d 4 to 6 in pregnant ewes. This decrease in concentrations of P4 from d 4 to 6 for CIDR-treated ewes likely indicates that a greater proportion of CIDR-treated ewes were either in anestrous, proestrus, or estrus compared with ewes not receiving a CIDR. The reduced concentrations of P4 compliment the difference found in that CIDR-treated had reduced d to estrus compared with ewes not receiving a CIDR. Similar to Bartlewski et al. (1999) who reported concentrations of P4 begin to increase between 3 and 7 7
CIDR-treated
6
Progesterone, ng/mL
Untreatd
d after estrus and then reach peak concentrations at approximately d 12, we found in the current study that CIDR-treated ewes when compared with untreated ewes follow this fluctuation of increased concentrations of P4 rising after d 6. However, peak levels of concentrations of P4 are seen at approximately d 20 for those ewes who were determined to be pregnant. Seasonal, lactational, and prepubertal ewes that are isolated from rams for at least a month, will exhibit a synchronized heat after re-introduction to the male approximately one to one and a half estrous cycles (17–24 d) later (Underwood et al., 1944; Martin et al., 1986). This has been termed the ‘ram effect’ (Underwood et al., 1944; Martin et al., 1986). It appears untreated ewes that become pregnant in the first service period were likely influenced by the ram effect which is evident by a delayed increase in concentrations of P4 compared with CIDR-treated ewes. This interval was relatively similar to that reported by Schinckel (1954), who indicated an interval of 14 d after ram introduction. Experiment 2: During the anestrous period (April through May) in which this study was conducted, CT and
trt*time
P = 0.36
5 4 3 2 1 0
-12
-5
0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 Day
Fig. 4. Serum progesterone (P4) profiles before and after 5-d controlled internal drug release (CIDR) synchronization protocols administered during the anestrous season of non-pregnant ewes (n = 46) in Experiment 2. Treatments: untreated = untreated; all CIDR based treatments = CIDR-treated.
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CTG ewes came into estrus more quickly after ram introduction than the CTP and CON ewes. This data indicates that CT and CTG treated ewes displayed a tighter synchrony to estrus. Similarly, Wheaton et al. (1992) observed ewes treated with a CIDR having a reduced interval to estrus compared with untreated ewes. In the current study, no differences were detected in overall days to lambing among treatment groups as well as all CIDR-treated ewes compared with CON. Conversely, Knights et al. (2001a) reported 42% of anestrous ewes treated with a 5 d CIDR lambed approximately 12–13 d earlier than untreated ewes. Similar to Knights et al. (2001a) and Wheaton et al. (1993) reported ewes in anestrous who were treated with a CIDR for 12 d required fewer d (157 ± 2 d) to lamb after introduction of rams compared with control ewes (170 ± 2 d). No statistical differences were detected among treatments for percentage of ewes exhibiting estrus to the first service and overall in the current study which is consistent with Wheaton et al. (1992) who reported no differences in ewes overall exhibiting estrus between untreated ewes and those receiving a CIDR insert for 12 d. Although we attempted to get all ewes in the anestrous period, prior to imposing treatments, ewes in the untreated group were already cycling as evidence by progesterone concentrations ≥1.0 ng/mL. Therefore, it is not surprising that pregnancy rates within the first service period after CIDR removal were similar in the current study for all treatments. In contrast, Knights et al. (2001a,b) documented greater pregnancy rates and an increase in lambing rate (Knights et al., 2001a) in P4 treated ewes compared with control ewes. In contrast to the current study, Titi et al. (2010) reported an increase in prolificacy in ewes treated with exogenous P4 when coupled with the use of GnRH and PG. Although no differences were detected for overall reproductive performance data, this study suggests that the CTG treatment was the most effective at concentrating estrus shortly after CIDR removal compared with CON, CT, and CTP treatments. In the present study, CIDR-treated ewes that were determined pregnant to the first service period had reduced concentrations of P4 from d 0 to 5. Perhaps this reduction in concentrations of P4 indicates decreased luteal activity which led to a more uniform synchronization to estrus further into the estrous cycle. Cyclicity during the anestrous season is not uncommon for the Dorset and Katahdin breeds as indicated by concentrations of P4 ≥ 1.0 ng/mL in 62% of the CON ewes; however, the overall pregnancy rate of 42% for CON ewes indicates CON ewes were not fully cyclic. It has been documented that a semi-cyclic pattern in ovarian activity is evident during early and late anestrous ewes (Cole and Miller, 1935). In the non-pregnant ewes, ewes had concentrations of P4 below 2.0 ng/mL from d 2 to 8. Both untreated and CIDR-treated ewes may have been in such a deep anestrous state that the treatments and/or ram effect were not powerful enough to bring them out of anestrous. Perhaps it did for a short period, explaining why the CIDR-treated ewes experienced an increase in concentrations of P4 around d 10; however, reverting back to decreased concentrations of P4 by d 15. Likewise, with the untreated ewes, an increase and then steady decline in concentrations of P4 can be observed from d
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1 to 14, with concentrations of P4 increasing again at d 15. Perhaps ram introduction stimulated cyclicity in the untreated ewes. These results are consistent with Martin et al. (1986), who reported that after 6 d of the short cycle, a second ovulation is induced followed by a fertile cycle of normal length. This apparent effect of ram introduction provides justification that a majority of the ewes were either non-cyclic or semi-cyclic. 5. Conclusions In the current study, during the transition and anestrous season 5 d CIDR inserts decreased overall days to detected estrus. Ewe CIDR-treatments decreased days to lambing in ewes bred during August and September. Regardless of season, inclusion of GnRH and PG into 5 d CIDR protocols did not improve ewe reproductive performance. The 5 d CIDR, regardless of protocol, did not markedly influence measures of reproductive performance desired by commercial sheep producers. The efficacy of CIDR protocols to synchronize estrus and improve fertility during the transition or anestrous period was not clearly demonstrated in this study. Conflict of interest None declared. Acknowledgements This study was supported by Zoetis Animal Health and the North Dakota State Board of Agriculture Research and Education Grant # 11-28. We thank the NDSU Sheep Shepherd Skip Anderson for all of his assistance during these studies. References Bartlewski, P.M., Beard, A.P., Rawlings, N.C., 1999. An ultrasonographic study of luteal function in breeds of sheep with different ovulation rates. Theriogenology 52, 115–130. Carlson, K.M., Pohl, H.A., Marcek, J.M., Muser, R.K., Wheaton, J.E., 1989. Evaluation of progesterone controlled internal drug release dispensers for synchronization of estrus in sheep. Anim. Reprod. Sci. 18, 205–218. Cole, H.H., Miller, R.F., 1935. Changes in the reproductive organs of the ewe with some data bearing on their control. Am. J. Anat. 57, 39–97. Glimp, H.A., 1971. Effect of breed and mating season on reproductive performance of sheep. J. Anim. Sci. 32, 1176–1182. Hamra, A.H., Massri, Y.G., Marcek, J.M., Wheaton, J.E., 1986. Plasma progesterone levels in ewes treated with progesterone-controlled internal drug-release dispensers, implants and sponges. Anim. Reprod. Sci. 11, 187–194. Hansel, W., Convey, E.M., 1983. Physiology of the estrous cycle. J. Anim. Sci. 57, 404–424. Knights, M., Hoehn, T., Lewis, P.E., Inskeep, E.K., 2001a. Effectiveness of intravaginal progesterone inserts and FSH for inducing synchronized estrus and increasing lambing rate in anestrous ewes. J. Anim. Sci. 79, 1120–1131. Knights, M., Maze, T.D., Bridges, P.J., Lewis, P.E., Inskeep, E.K., 2001b. Shortterm treatment with a controlled internal drug releasing (CIDR) device and FSH to induce fertile estrus and increase prolificacy in anestrous ewes. Theriogenology 55, 1181–1191. Martin, G.B., Oldham, C.M., Cognie, Y., Pearce, D.T., 1986. The physiological responses of anovulatory ewes to the introduction of rams – a review. Livest. Prod. Sci. 15, 219–247. Schinckel, P.G., 1954. The effect of the presence of the ram on the ovarian activity of the ewe. Aust. J. Agric. Res. 5, 465–469.
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Schneider, F.A., Hallford, D.M., 1996. Use of a rapid progesterone radioimmunoassay to predict pregnancy and fetal numbers in ewes. Sheep Goat Res. J. 12, 33–38. Titi, H.H., Kridli, R.T., Alnimer, M.A., 2010. Estrus synchronization in sheep and goats using combinations of GnRH, progestagen and prostaglandin F2␣ . Reprod. Domest. Anim. 45, 594–599. Underwood, E.J., Shier, F.L., Davenport, N., 1944. Studies in sheep husbandry in W.A. V. The breeding season of Merino, Crossbred and British breed ewes in the agricultural districts. J. Agric. West. Aust. 2, 135–143.
Wheaton, J.E., Windels, H.F., Johnston, L.J., 1992. Accelerated lambing using exogenous progesterone and the ram effect. J. Anim. Sci. 70, 2628–2635. Wheaton, J.E., Carlson, K.M., Windels, H.F., Johnston, L.J., 1993. CIDR: a new progesterone-releasing intravaginal device for induction of estrus and cycle control in sheep and goats. Anim. Reprod. Sci. 33, 127–141. Wildeus, S., 2000. Current concepts in synchronization of estrus: sheep and goats. J. Anim. Sci. 77, 1–14.
Please cite this article in press as: Jackson, C.G., et al., Efficacy of various five-day estrous synchronization protocols in sheep. Small Ruminant Res. (2014), http://dx.doi.org/10.1016/j.smallrumres.2014.04.004