Pregnancy per artificial insemination in lactating dairy cows subjected to 2 different intervals from presynchronization to initiation of Ovsynch protocol

A protocol for presynchronization of ovarian status with 2 injections of PGF_2α given 14 d apart, with the last PGF_2α injection given 12 or 14 d before Ovsynch increases pregnancy per artificial insemination (P/AI) in dairy cows. We determined the efficacy of reducing the interval from the last PGF_2α injection (500 μg of cloprostenol) of presynchronization to initiation of Ovsynch on response to treatment and P/AI. Lactating dairy cows were assigned to an Ovsynch protocol, with the initial injection of GnRH given either 9 (PRE-9; n = 135) or 12 d (PRE-12; n = 135) after the second PGF_2α injection of presynchronization. The Ovsynch protocol consisted of 2 injections of 100 μg of GnRH given 9 d apart and 1 injection of PGF_2α given 7 d after the initial GnRH injection, and cows were subjected to timed artificial insemination (TAI; 70 ± 3.5 DIM) approximately 16 h after the second GnRH injection. Body condition score (1–5 scale) was recorded at TAI. Blood samples were taken for progesterone determination at the PGF_2α injection of Ovsynch, at TAI, and at 11 d after TAI. Ultrasonographic examinations were done in all cows at the second PGF_2α injection of presynchronization, initial GnRH injection, PGF_2α injection of Ovsynch, at TAI, and 24 h after TAI for cyclicity status and ovarian responses to treatments, and at 32 and 60 d after TAI for confirmation of pregnancy. Overall, 29 cows (10.7%) were determined acyclic or cystic and excluded from the study. The percentage of cows responding to initial GnRH injection (62.2 vs. 61.5%) did not differ between PRE-9 and PRE-12 but more cows in the PRE-9 group failed to respond to PGF_2α treatment of Ovsynch compared with PRE-12 (22.7 vs. 10.7%). Body condition score at TAI (2.9 ± 0.02) and mean ovulatory follicle diameter (16.4 ± 0.2 mm) were not different between treatments. Overall P/AI at 32 d was reduced in PRE-9 (33.6%) compared with PRE-12 (44.3%) but pregnancy losses (5.0 vs. 3.7%) did not differ between treatments. Primiparous cows in the PRE-12 group had higher mean progesterone concentration 11 d after TAI and greater P/AI 32 after TAI than primiparous cows in the PRE-9 group (6.4 ± 0.5 vs. 4.6 ± 0.5 ng/mL and 55.8 vs. 30.0%, respectively). In conclusion, reducing the interval from the last PGF_2α injection of the presynchronization treatment to initiation of Ovsynch (from 12 to 9 d) did not affect ovulatory response to initial GnRH injection but reduced response to PGF_2α injection of Ovsynch and P/AI at 32 and 60 d after TAI. The reduction in P/AI was particularly evident in primiparous cows of the PRE-9 group.     

Presynchronization using a modified Ovsynch protocol or a single gonadotropin-releasing hormone injection 7 d before an Ovsynch-56 protocol for submission of lactating dairy cows to first timed artificial insemination

Presynchronization strategies, such as Presynch-Ovsynch and Double-Ovsynch, increase fertility to timed artificial insemination (TAI) compared with Ovsynch alone; however, simpler presynchronization strategies could reduce costs and simplify reproductive management. Lactating Holstein cows (n = 601) were randomly assigned to 1 of 2 presynchronization treatments before beginning an Ovsynch-56 protocol (GnRH at 70 ± 3 DIM, PGF_2α 7 d later, GnRH 56 h after PGF_2α, and TAI 16 h later at 80 ± 3 DIM) for first TAI. Cows (n = 306) in the first treatment (Double-Ovsynch; DO) were presynchronized using a modified Ovsynch protocol (GnRH at 53 ± 3 DIM, 7 d later PGF_2α, and GnRH 3 d later) ending 7 d before the first GnRH injection (G1) of an Ovsynch-56 protocol. Cows (n = 295) in the second treatment (GGPG) were presynchronized using a single injection of GnRH 7 d before G1 of an Ovsynch-56 protocol at 63 ± 3 DIM. Blood samples were collected at G1 and the PGF_2α injections of the Ovsynch-56 protocol to determine progesterone (P4) concentrations. Pregnancy diagnosis was performed using ultrasonography 32 d after TAI, and pregnant cows were reexamined 46 and 70 d after TAI. Overall, DO cows had more pregnancies per artificial insemination (P/AI) compared with GGPG cows 32 d after TAI (53 vs. 43%). Overall, P/AI did not differ by parity (primiparous vs. multiparous), and pregnancy loss did not differ between treatments or parities. More DO cows had P4 in a medium range (>0.5 to <4 ng/mL) at G1 of the Ovsynch-56 protocol compared with GGPG cows (82 vs. 50%), and more DO cows had high P4 (>4 ng/mL) at the PGF_2α injection of the Ovsynch-56 protocol compared with GGPG cows (67 vs. 36%). Thus, presynchronization with a modified Ovsynch protocol increased P/AI after TAI at first AI by increasing synchrony to the Ovsynch-56 protocol compared with presynchronization using a single injection of GnRH.     

Effect of pretreatment with prostaglandin F2alpha before resynchronization of ovulation on fertility of lactating dairy cows

Our objective was to assess the effect of pretreatment with PGF_2α 12 d before initiation of a protocol for resynchronization of ovulation (Resynch) using an Ovsynch protocol. Lactating Holstein cows diagnosed not pregnant 31 d after a timed artificial insemination (TAI) were randomly assigned to initiate the Resynch protocol 32 d after TAI (n = 255; RES), or receive 25 mg of PGF_2α 34 d after TAI and initiate the Resynch protocol 12 d later at 46 d after TAI (n = 272; PGF+RES). Within each treatment, a subset of cows were examined using transrectal ultrasonography to determine ovulatory response to the first GnRH injection of the Resynch protocols or a blood sample was collected to determine serum progesterone (P₄) at initiation of the Resynch protocol, or both. Overall, PGF+RES cows had more pregnancies per artificial insemination (P/AI) than RES cows 66 d after TAI (35.2 vs. 25.6%), whereas pregnancy loss from 31 to 66 d after TAI was greater for RES than PGF+RES cows (17.1 vs. 7.6%). Although P/AI was greater for cows with high (≥1.0 ng/mL) vs. low (<1.0 ng/mL) P₄ at the first GnRH injection of the Resynch protocols, treatment did not affect the proportion of cows with low P₄ at the first GnRH injection of the Resynch protocols. Overall, no effect of treatment on ovulatory response to the first GnRH injection of the Resynch protocols was detected. We conclude that pretreatment with PGF_2α 12 d before initiation of the Resynch protocol increased P/AI 66 d after TAI for cows with serum P₄ concentration >1.0 ng/mL at the first GnRH injection of the Resynch protocol and decreased pregnancy loss from 31 to 66 d after TAI. This modified resynchronization protocol may be a useful strategy for reproductive management of lactating dairy cows.     

Effect of dose and timing of prostaglandin F2α treatments during a Resynch protocol on luteal regression and fertility to timed artificial insemination in lactating Holstein cows

Our objective was to evaluate the effect of a second PGF_2α treatment (25 mg of dinoprost) or a double dose of PGF_2α (50 mg of dinoprost) during a Resynch protocol on luteal regression and pregnancies per artificial insemination (P/AI) in lactating dairy cows. Lactating Holstein cows (n = 1,100) were randomly assigned at a nonpregnancy diagnosis to receive (1) Ovsynch (control: 100 µg of GnRH; 7 d, 25 mg of PGF_2α; 56 h, 100 µg of GnRH), (2) Ovsynch with a second PGF_2α treatment (GPPG: 100 µg of GnRH; 7 d, 25 mg of PGF_2α; 24 h, 25 mg of PGF_2α; 32 h, 100 µg of GnRH), or (3) Ovsynch with a double dose of PGF_2α (GDDP: 100 µg of GnRH; 7 d, 50 mg of PGF_2α; 56 h, 100 µg of GnRH). All cows received timed artificial insemination (TAI) approximately 16 h after the second GnRH treatment (G2). Pregnancy diagnosis was performed by transrectal palpation 39 ± 3 d after TAI, and pregnancy status was reconfirmed 66 d after TAI. Blood samples collected from a subset of cows in each treatment at the first PGF_2α treatment (n = 394) and at G2 (n = 367) were assayed for progesterone (P4). Data were analyzed by logistic regression using the GLIMMIX procedure of SAS (SAS Institute Inc., Cary, NC). At 39 d after TAI, GPPG cows tended to have more P/AI than control cows [35% (137/387) vs. 31% (107/349)], whereas P/AI for GDDP cows [32% (118/364)] did not differ from that for control cows. Pregnancy loss from 38 to 66 d did not differ among treatments and was 8% (30/362). The percentage of cows with complete luteal regression (P4 <0.4 ng/mL at G2) tended to differ among treatments and was greater for GPPG cows than for GDDP and control cows (94% vs. 88% vs. 88%, respectively). Overall, cows with P4 <1 ng/mL at the first PGF_2α treatment had fewer P/AI than cows with P4 ≥1 ng/mL (27% vs. 38%), whereas cows with P4 ≥0.4 ng/mL at G2 had fewer P/AI than cows with P4 <0.4 ng/mL (15% vs. 38%). We conclude that adding a second PGF_2α treatment 24 h after the first within a Resynch protocol tended to increase the proportion of cows undergoing complete luteal regression and P/AI, whereas treatment with a double dose of PGF_2α at a single time did not.     

Detection of anovulation by heatmount detectors and transrectal ultrasonography before treatment with progesterone in a timed insemination protocol

Our objective was to determine the accuracy of identifying noncycling lactating dairy cows before the application of a timed artificial insemination (AI) protocol [with or without progesterone supplementation via a controlled internal drug-release (CIDR) insert and 2 different timings of AI] by using heatmount detectors and a single ovarian ultrasound examination. At 6 locations in the Midwest, 1,072 cows were enrolled in a Presynch protocol (2 injections of PGF_2α 14 d apart), with the second injection administered 14 d before initiating the Ovsynch protocol (injection of GnRH 7 d before and 48 h after PGF_2α injection, with timed AI at 0 or 24 h after the second GnRH injection). Heatmount detectors were applied to cows just before the first Presynch injection, assessed 14 d later at the second Presynch injection (replaced when activated or missing), and reassessed at initiation of the Ovsynch protocol. Ovaries were examined for the presence of a corpus luteum (CL) by ultrasound before the initiation of treatment. Treatments were assigned to cows based on the presence or absence of a CL detected by ultrasound: 1) no CL + no CIDR; 2) no CL + CIDR insert for 7 d; and 3) CL present. Further, alternate cows within the 3 treatments were assigned to be inseminated concurrent with the second GnRH injection of Ovsynch (0 h) or 24 h later. Pregnancy was diagnosed at 33 and 61 d after the second GnRH injection. By using low (<1 ng/mL) concentrations of progesterone in serum as the standard for noncycling status, heatmount detectors were activated on a large percentage of noncycling cows (>60%), whereas the single ultrasound examination incorrectly classified noncycling cows only 21% of the time. Conversely, cycling cows (progesterone ≥1 ng/mL) were correctly identified 70 to 78% of the time by heatmount detectors, but 85 to 92% were correctly identified by ultrasound. Overall accuracy of heatmount detectors and ultrasound was 71 and 84%, respectively. Application of progesterone to cows without a CL at the time of the first injection of GnRH reduced the incidence of ovulation but increased the proportions of pregnancies per AI at d 33 or 61 compared with nontreated cows without a CL at the onset of the Ovsynch protocol. Percentages of cows pregnant and pregnancy survival did not differ for cows having a CL before treatment compared with those not having a CL and treated with progesterone. Compared with no response, when a follicle ovulated in response to the first GnRH injection, percentage of cows becoming pregnant after the timed AI increased from 33.3 to 41.6%. Timing of AI at 0 or 24 h after the second GnRH injection did not alter pregnancies per AI, but cows having luteal activity before treatment had improved pregnancies per AI compared with noncycling cows. We conclude that identifying noncycling cows by ultrasound was more accurate than by heatmount detectors. Subsequent progesterone treatment of previously cycling cows not having a CL at the onset of Ovsynch increased the proportion of pregnant cows, equal to that of cows having a CL but not treated with progesterone.     

Optimizing ovulation to first GnRH improved outcomes to each hormonal injection of ovsynch in lactating dairy cows

Ovulatory response to the first GnRH of Ovsynch is the critical determinant for successful synchronization of ovulation in dairy cows. Our objective in this study was to develop a pre-Ovsynch treatment that increased the percentage of cows that ovulated in response to the first GnRH injection of Ovsynch. To accomplish our goal, we evaluated a hormonal strategy that consisted of PGF_2α and GnRH before the first GnRH of Ovsynch. Lactating dairy cows (n = 137) were assigned to receive either no treatment before Ovsynch (control) or 25 mg of PGF_2α (PreP) followed 2 d later by 100 μg of GnRH (PreG), administered 4 (G4G), 5 (G5G), or 6 (G6G) d before initiating the Ovsynch protocol. Transrectal ultrasonography was performed to assess follicular size and resulting ovulation, and blood samples were collected to measure circulating concentrations of progesterone and estradiol immediately before each hormonal injection. Cows were inseminated at a fixed time 16 h after final GnRH of Ovsynch. Pregnancy diagnosis was performed 35 d later by palpation per rectum of uterine contents. Proportion of cows that ovulated to first GnRH of Ovsynch was 56.0, 66.7, 84.6, and 53.8% for G4G, G5G, G6G, and controls, respectively, and was greater for G6G than for control cows. Luteolytic response to PGF_2α of Ovsynch was greater in all treated than control cows (92.0, 91.7, 96.2, and 69.2% for G4G, G5G, G6G, and control, respectively). Synchronization rate to Ovsynch was greater (92 vs. 69%, respectively) in G6G than in control cows. In addition, cows that ovulated in response to first GnRH of Ovsynch had greater response to PGF_2α of Ovsynch (92.7 vs. 77.1%, respectively) and greater synchronization rate to the overall protocol (87.9 vs. 62.9%, respectively) than those that did not ovulate. Concentrations of progesterone at PGF_2α of Ovsynch, and estradiol and follicle size at final GnRH of Ovsynch, were identified as significant predictors of probability of pregnancy 35 d after artificial insemination. In summary, a PGF_2α-and-GnRH based pre-Ovsynch strategy consisting of a 6-d interval between PreG and first GnRH of Ovsynch resulted in a greater ovulatory and luteolytic response to first GnRH and PGF_2α of Ovsynch, respectively, compared with control cows. This, in turn, optimized synchronization rate to Ovsynch.     

Effect of interval to resynchronization of ovulation on fertility of lactating Holstein cows when using transrectal ultrasonography or a pregnancy-associated glycoprotein enzyme-linked immunosorbent assay to diagnose pregnancy status

The objective of this study was to compare 2 strategies for resynchronization of ovulation based on nonpregnant diagnoses using transrectal ultrasonography or a pregnancy-associated glycoprotein (PAG) ELISA. Lactating Holstein cows (n = 1,038) were submitted for first postpartum timed artificial insemination (TAI) using a Presynch + Ovsynch protocol. After the initial breeding, cows were randomly assigned to initiate resynchronization 25 d (D25) or 32 d (D32) later. Pregnancy status of cows initiating Resynch 25 d after TAI was determined 27 d after TAI by using a PAG ELISA, whereas pregnancy status of cows initiating Resynch 32 d after TAI was determined 39 d after TAI using transrectal ultrasonography. Cows diagnosed as not pregnant continued the Resynch protocol by receiving an injection of PGF_2α 7 d after the initial GnRH injection and a second GnRH injection 54 h after the PGF_2α injection. Cows in both treatments were inseminated approximately 16 h after the second GnRH injection. Blood samples for analysis of progesterone (P₄) were collected at the first GnRH injection of each Resynch protocol. Pregnancies per AI (P/AI) of nonpregnant cows initiating Resynch 25 vs. 32 d after first postpartum TAI did not differ 39 d after TAI and were 28.3 vs. 30.9% for D25 vs. D32 cows, respectively. Mean P₄ at the first GnRH injection of Resynch was greater for D32 than for D25 cows (3.67 ± 0.22 vs. 2.83 ± 0.22 ng/mL), indicating that the Resynch treatments were initiated at different stages of the estrous cycle. After blocking P₄ concentration into low (<1.0 ng/mL) or high (≥1.0 ng/mL) classes, P₄ class was not found to affect P/AI 39 d after TAI. Early resynchronization was not found to affect P/AI 39 d after TAI; however, early resynchronization did decrease days between inseminations and the interval from the initial nonpregnant diagnosis to conception. Earlier detection of nonpregnant cows using the PAG ELISA in conjunction with a TAI resynchronization program may improve the rate at which cows become pregnant in a dairy herd compared with transrectal ultrasonography conducted at a later stage after TAI.     

The effect of presynchronization with prostaglandin F2α and gonadotropin-releasing hormone simultaneously, 7 d before Ovsynch,compared with Presynch-10/Ovsynch on luteal function and first-service pregnancies per artificial insemination

Pregnancy per artificial insemination (P/AI) following Ovsynch is optimized when cows ovulate to the first GnRH of Ovsynch. Fertility programs are designed to presynchronize cows to d 6 or 7 of the estrous cycle to increase the chances of ovulation of a first-wave dominant follicle to the first GnRH of Ovsynch. The hypothesis of this experiment was that simplification of a presynchronization program through the combination of PGF_2α and GnRH on the same day, 7 d before Ovsynch, would allow for similar P/AI compared with Presynch-10. Lactating dairy cows (n = 432) 41 to 47 d in milk (DIM) were randomly assigned to 2 treatments within parities for first service. Control cows received Presynch-10/Ovsynch consisting of the following: PGF_2α–14 d–PGF_2α–10 d–GnRH–7 d–PGF_2α–56 h–GnRH–16 h–AI. Treated cows received PGF_2α and GnRH–7 d–GnRH–7 d–PGF_2α–56 h–GnRH–16 h–AI. All cows received a supplemental injection of PGF_2α 24 h after the PGF_2α of Ovsynch to enhance complete luteolysis. All cows received timed AI between 75 and 81 DIM. Blood was collected to assess circulating concentrations of progesterone (P4), and the number and size of corpora lutea (CL) were recorded using ultrasonography on day of PGF_2α of Ovsynch. The administration of PGF_2α simultaneously with GnRH and 7 d before Ovsynch (PG+G) had similar P/AI at 28 (46 vs. 48%), 35 (43 vs. 43%), 49 (39 vs. 39%), and 77 d post-AI (38 vs. 39%) compared with Presynch-10. No differences were observed in P/AI in primiparous versus multiparous cows at 28 (52 vs. 45%), 35 (48 vs. 41%), 49 (45 vs. 37%), and 77 d post-AI (43 vs. 36%). No difference existed between treatments in percentage of cows with functional CL at PGF_2α of Ovsynch, total luteal area (mm²), or serum concentrations of P4 at time of PGF_2α of Ovsynch, regardless of parity. Number of CL had a tendency to be greater for multiparous PG+G vs. Presynch-10 cows (2.34 ± 0.09 vs. 2.15 ± 0.08) but not in primiparous cows (1.95 ± 0.10 vs. 1.98 ± 0.11). In summary, administering both PGF_2α and GnRH on the same day, 7 d before the start of Ovsynch, appears to be a simple and effective alternative to Presynch-10 Ovsynch.     

Pregnancy rates to timed artificial insemination in Holstein heifers given prostaglandin F2alpha twenty-four hours before or concurrent with removal of an intravaginal progesterone-releasing insert

The objective was to compare pregnancy rates in nulliparous Holstein heifers given PGF_2α 24 h before, or concurrent with, removal of an intravaginal progesterone-releasing (CIDR) insert in 3 timed artificial insemination (TAI) protocols. Heifers (from 2 herds) were assigned randomly, over 11 mo, to 1 of 3 modified Ovsynch protocols. On d 0 (without reference to the stage of the estrous cycle), all heifers were given 100 μg of GnRH i.m. and a CIDR insert (containing 1.9 g of progesterone). In the PG-7/P4-8 protocol (n = 99), PGF_2α was given on d 7, and CIDR inserts were removed on d 8. In the PG-7/P4-7 (n = 98) and PG-8/P4-8 (n = 102) protocols, PGF_2α administration and CIDR removal occurred concurrently, on d 7 or 8, respectively. In all 3 protocols, a second GnRH treatment (100 μg) was given 48 h after PGF_2α with TAI 16 to 20 h later. Blood samples were collected (subset of 124 heifers) on d 0, 7, 10 or 11 (i.e., at TAI), and 17. Pregnancy rates (32 d after TAI) for protocols PG-7/P4-8, PG-7/P4-7, and PG-8/P4-8 were 61.8, 55.6, and 54.1%, respectively. Pregnancy rate was higher when synchronization was initiated during diestrus than when initiated at other stages (57.0 versus 34.8%). Although pregnancy rates were not affected by season, there was an interaction between protocol and season; pregnancy rates were significantly lower in summer in heifers subjected to PG-7/P4-7 and PG-8/P4-8, but season did not affect pregnancy rates in heifers subjected to PG-7/P4-8. In summary, giving PGF_2α 24 h before CIDR removal, followed by TAI (PG-7/P4-8 protocol), resulted in consistent pregnancy rates, regardless of season, relative to protocols involving PGF_2α treatment concurrent with CIDR removal.     

Supplementation of progesterone via controlled internal drug release inserts during ovulation synchronization protocols in lactating dairy cows

Our objective was to determine the effect of exogenous progesterone (P4) during a timed artificial insemination (TAI) protocol on pregnancies per AI (P/AI) in dairy cows not previously detected in estrus. Lactating cows (n = 3,248) from 7 commercial dairy herds were submitted to a presynchronization protocol (2 injections of PGF_2α 14 d apart; Presynch), and cows in estrus after the second PGF_2α received AI (EDAI; n = 1,583). Cows not inseminated by 12 to 14 d after the second PGF_2α injection were submitted to a TAI protocol (GnRH on d 0, PGF_2α on d 7, and GnRH + TAI 72 h after PGF_2α). At onset of the TAI protocol, cows were balanced by parity and days in milk and assigned randomly to receive no exogenous P4 (control, n = 803) or a controlled internal drug release (CIDR) insert containing 1.38 g of P4 from d 0 to 7 (CIDR, n = 862). Blood samples were collected at the second PGF_2α injection of the Presynch and on the day of the first GnRH injection of the TAI protocol for P4 determination. When P4 in both samples was <1 ng/mL, cows were classified as anovular, whereas cows having at least 1 sample ≥1 ng/mL were classified as cyclic. Concentration of P4 at 11 to 14 d after AI was determined in a subgroup of cows (n = 453) from 2 herds. Pregnancy was diagnosed at 40 ± 5 and 65 ± 5 d after AI. Proportion of cows inseminated on estrus after the second PGF_2α injection of the Presynch protocol differed among herds (range = 26.7 to 59.8%). Overall P/AI for EDAI cows at 40 ± 5 and 65 ± 5 d were 36.2 and 33.7%, respectively, and pregnancy loss was 8.8%. Proportion of cyclic cows at the onset of the TAI protocol differed among herds (range from 66.5 to 86.3%), but did not differ between treatments (control = 72.4%, CIDR = 74.1%). Treatment affected P/AI at 40 ± 5 (control = 33.3%, CIDR = 38.1%) and 65 ± 5 (control = 30.0%, CIDR = 35.1%) d after AI but did not affect pregnancy loss (8.6%). Cyclic cows had greater P/AI at 40 ± 5 (38.2 vs. 29.3%) and 65 ± 5 d (35.1 vs. 26.1%) after AI, but cyclic status had no effect on pregnancy loss. Treatment affected P4 concentration after AI, with more CIDR cows having P4 ≥1 ng/mL (94.4 vs. 86.9%) and P4 ≥3.2 ng/mL (81.8 vs. 68.0%) at 11 to 14 d after AI compared with control cows. Treatment of cows not previously detected in estrus with a CIDR insert during a TAI protocol increased proportion of cows with functional CL after AI and P/AI.     

Alternative programs to presynchronize estrous cycles in dairy cattle before a timed artificial insemination program

The objective was to test potential presynchronization programs applied to cows before a timed artificial insemination (TAI) program to increase the percentage of cows ovulating in response to both GnRH injections of a TAI program and having a functional corpus luteum before the first GnRH injection of the TAI program. At calving, cows were blocked by lactation (1 vs. 2+) and assigned randomly to receive 1 of 5 presynchronization treatments. Two variants of the standard Presynch program were tested in which 2 injections of PGF_2α were administered 14 d apart with either 14 d (Pre14; n = 122), 12 d (Pre12; n = 123), or 10 d (Pre10; n = 151) intervening before a TAI program was initiated. Two other presynchronization programs consisted of administering a progesterone-releasing controlled internal drug release (CIDR) insert for 7 d plus PGF_2α administration at insert removal. Insert removal occurred either 10 d (CIDR10; n = 157) or 3 d (CIDR3; n = 117) before a TAI program was initiated. The TAI program was a standard Cosynch program [injection of GnRH 7 d before (GnRH-1) and 72 h after (GnRH-2) PGF_2α with TAI administered 72 h after PGF_2α). Cosynch served as the control (n = 157), and cows were assumed to be starting this program at random stages of the estrous cycle. From a subset of cows per treatment (ranging from 49 to 51 cows each), blood samples were collected from coccygeal vessels by using evacuated tubes at d −28, −14, 0 (onset of TAI program), 7, 9, 14, and 21. Ovarian scans were conducted on d 0, 7, 9, 14, and 21 by transrectal ultrasonography. Diameters of follicles and corpus luteum were measured at each exam, and ovulation was determined on d 7 (response to GnRH-1 on d 0) and d 14 (response to GnRH-2 on d 10). Ovulatory incidence after GnRH-1 (47.1 to 67.3%) and GnRH-2 (78 to 90.2%) varied but did not differ among treatments. Before GnRH-1, progesterone concentrations were less in the CIDR3 treatment than in all other treatments. Before GnRH-2, progesterone was greater in the CIDR3 treatment than in all other treatments. Luteal regression and synchronization rate (successful luteolysis and ovulation after GnRH-2) did not differ among treatments. Pregnancy rate per AI at 32 and 60 d post TAI was less in CIDR3 cows than in cows in all other treatments. None of the Presynch treatments improved key responses (ovulation, luteolysis, and synchronization rate) known to improve fertility compared with a standard Cosynch program without presynchronization.     

Effect of increasing amounts of supplemental progesterone in a timed artificial insemination protocol on fertility of lactating dairy cows

The objectives were to evaluate the effect of supplemental progesterone during a timed artificial insemination (TAI) protocol on pregnancy per insemination and pregnancy loss. Lactating dairy cows from 2 dairy herds were presynchronized with 2 injections of PGF_2α 14 d apart, and cows observed in estrus following the second PGF_2α injection were inseminated (n = 1,301). Cows not inseminated by 11 d after the end of the presynchronization were submitted to the TAI protocol (d 0 GnRH, d 7 PGF_2α, d 8 estradiol cypionate, and d 10 TAI). On the day of the GnRH of the TAI protocol (study d 0), cows were assigned randomly to receive no exogenous progesterone (control = 432), one controlled internal drug-release (CIDR) insert (CIDR1 = 440), or 2 CIDR inserts (CIDR2 = 440) containing 1.38 g of progesterone each from study d 0 to 7. Blood was sampled on study d 0 before insertion of CIDR for determination of progesterone concentration in plasma, and cows with concentration <1.0 ng/mL were classified as low progesterone (LP) and those with concentration ≥1.0 ng/mL were classified as high progesterone (HP). From a subgroup of 240 cows, blood was sampled on study d 3, 7, 17 and 24 and ovaries were examined by ultrasonography on study d 0 and 7. Pregnancy was diagnosed at 38 ± 3 and 66 ± 3 d after AI. Data were analyzed including only cows randomly assigned to treatments and excluding cows that were inseminated after the second PGF_2α injection. The proportion of cows classified as HP at the beginning of the TAI protocol was similar among treatments, but differed between herds. Concentrations of progesterone in plasma during the TAI protocol increased linearly with number of CIDR used, and the increment was 0.9 ng/mL per CIDR. The proportion of cows with plasma progesterone ≥1.0 ng/mL on study d 17 was not affected by treatment, but a greater proportion of control than CIDR-treated cows had asynchronous estrous cycles following the TAI protocol. Treatment with CIDR inserts, however, did not affect pregnancy at 38 ± 3 and 66 ± 3 d after AI or pregnancy loss.     

Assessment of a practical method for identifying anovular dairy cows synchronized for first postpartum timed artificial insemination

Our objective was to determine whether a single examination of ovaries using transrectal ultrasonography at the first GnRH injection of a Presynch + Ovsynch protocol is a useful method for assessing cyclicity status and thereby enabling differential management of anovular vs. cyclic cows. Lactating Holstein cows (n = 842) receiving a Presynch + Ovsynch protocol to initiate first postpartum timed artificial insemination (TAI) were used to compare 2 methods for assessing cyclicity status before TAI. For the standard method (using RIA), blood samples were collected at the second PGF_2α injection of Presynch and the first GnRH injection of Ovsynch, and cows with serum progesterone ≥1.0 ng/mL in one or both samples were classified as cycling, whereas cows with serum progesterone <1.0 ng/mL in both samples were classified as anovular. For the practical method, transrectal ultrasonography (U/S) was used to determine the presence or absence of a corpus luteum (CL) at the first GnRH injection of Ovsynch, and cows without CL were classified as anovular, whereas cows with CL were classified as cycling. Statistical agreement (kappa) between the RIA and U/S methods to identify cycling cows was 0.66. Sensitivity, specificity, positive predictive value, and negative predictive value of U/S to identify anovular status were 85.7, 87.7, 64.7, and 95.9%, respectively. We conclude that assessing the presence or absence of CL at the first GnRH injection of a Presynch + Ovsynch protocol using U/S is a reliable and practical method for identifying the cyclicity status of cows before first TAI, but may slightly overestimate the proportion of anovular cows compared with the RIA method.     

Timed artificial insemination of two consecutive services in dairy cows using prostaglandin F2alpha and gonadotropin-releasing hormone

Timed artificial insemination (TAI) protocols use PGF_2α and GnRH injections to synchronize ovulation. The objective was to evaluate the PGPG protocol (d 0, PGF_2α; d 3, GnRH; d 11, PGF_2α; d 13, GnRH and TAI) for first TAI and also examine methods for second TAI in nonpregnant cows. A factorial test of the first PGF_2α and first GnRH injections within the PGPG protocol was performed (the last PGF_2α and GnRH injections were deemed essential to the TAI). Lactating dairy cows (n = 804) in a commercial herd were assigned to 1 of 5 first-TAI treatments, which were PGPG, GPG (d 0, no treatment; d 3, GnRH; d 11, PGF_2α; d 13, GnRH and TAI), PPG (d 0, PGF_2α; d 3, no treatment; d 11, PGF_2α; d 13, GnRH and TAI), and PG (d 0, no treatment; d 3, no treatment; d 11, PGF_2α; d 13, GnRH and TAI); the Ovsynch protocol (GnRH, 7 d, PGF_2α, 2 d, GnRH and TAI) was the positive control. For resynchronization, cows received either GnRH or the control (no injection) on d 22 after TAI. Nonpregnant cows on d 28 were then treated with PGF_2α on d 29, GnRH on d 31, and TAI [i.e., resynchronization treatments of ReGPG (received GnRH on d 22) and RePG (did not receive GnRH on d 22)]. Pregnancy rates for PGPG, GPG, PPG, PG, and Ovsynch were similar at d 28 after first TAI. Analyses of multiple explanatory factors by logistic regression detected an effect of uterine or ovarian abnormality on the d-28 pregnancy rate (normal more likely to be pregnant). Day-42 pregnancy rates were affected by uterine or ovarian abnormality (normal more likely to be pregnant), postpartum disease occurrence (healthy cows more likely to be pregnant), milk production, and days in milk. Treatment was not significant for the d-42 pregnancy rate. Effects of postpartum disease, milk production, and days in milk on the d-42 pregnancy rate were apparently manifested through their effects on embryonic loss between d 28 and 42 of pregnancy. High-producing cows that received TAI early postpartum were most likely to experience embryonic loss. Day-42 pregnancy rates after the resynchronization treatment were affected by an interaction of the first synchronization treatment with the resynchronization treatment. We concluded that although PGPG can be used for TAI, a simpler TAI protocol that includes the last 2 injections (PGF_2α, 2 d; GnRH and TAI) would be equally effective.     

Comparison of 2 protocols to increase circulating progesterone concentration before timed artificial insemination in lactating dairy cows with or without elevated body temperature

Two treatments designed to increase circulating progesterone concentration (P4) during preovulatory follicle development were compared. One treatment used 2 intravaginal P4 implants (controlled internal drug-releasing inserts; CIDR) and the other used a GnRH treatment at beginning of the protocol. Lactating Holstein cows that had been diagnosed as nonpregnant were randomly assigned to receive timed artificial insemination (TAI) following 1 of 2 treatments (n = 1,638 breedings): (1) GnRH: CIDR+ 2 mg of estradiol (E2) benzoate + 100 µg of GnRH on d ?11, PGF_2α on d ?4, CIDR withdrawal + 1.0 mg of E2-cypionate + PGF_2α) on d ?2, and TAI on d 0; or (2) 2CIDR: 2 CIDR + 2 mg of E2-benzoate on d ?11, 1 CIDR withdrawn + PGF_2α on d ?4, second CIDR withdrawn + 1.0 mg of E2-cypionate + PGF_2α on d ?2, and TAI on d 0. Milk yield was measured daily between d 0 and d 7. Rectal temperature was measured using a digital thermometer at d 0 and 7, and elevated body temperature was defined as an average rectal temperature ≥39.1°C. Pregnancy diagnoses were performed on d 32 and 60 after TAI. We detected no effect of treatments on pregnancy per AI or pregnancy loss regardless of elevated body temperature, body condition score, parity, milk yield, or presence or absence of a corpus luteum (CL) on d ?11 or d ?4. Pregnancy per AI at 60 d was reduced [elevated body temperature = 22.8% (162/709), no elevated body temperature 34.1% (279/817)] and pregnancy loss tended to increase [elevated body temperature = 20.2% (41/203), no elevated body temperature 14.4% (47/326)] in cows with elevated body temperature. Various physiological measurements associated with greater fertility were also reduced in cows with elevated body temperature, such as percentage of cows with a CL at PGF_2α (decreased 7.9%), ovulatory follicle diameter (decreased 0.51 mm), expression of estrus (decreased 5.1%), and ovulation near TAI (decreased 2.8%) compared with cows without elevated body temperature. A greater proportion of cows (30.2%) had a CL at PGF_2α in the GnRH treatment [74.1% (570/763)] than in the 2CIDR treatment [56.9% (434/763)]; however, circulating P4 concentration was greater at the time of PGF_2α treatment (d ?4) for cows 2CIDR (4.26 ± 0.13 ng/mL) than in cows in GnRH (3.99 ± 0.14 ng/mL). Thus, these 2 protocols yield similar fertility results that might be due to somewhat different physiological alterations. Treatment with GnRH increased the proportion of cows with a CL at PGF_2α; however, the 2CIDR protocol increased circulating P4 under all circumstances.     

Comparison of reproductive performance in lactating dairy cows bred by natural service or timed artificial insemination

The objective of this study was to compare reproductive performance of lactating dairy cows bred by natural service (NS) or timed AI (TAI). One thousand fifty-five cows were blocked by parity and enrolled to receive either NS or TAI. Cows in both groups were presynchronized with 2 injections of PGF_2α given at 42 and 56 d postpartum. Fourteen days after the last PGF_2α injection, cows in the TAI group were enrolled in an Ovsynch protocol (d 0 GnRH; 7 d later, PGF_2α; 56 h after PGF_2α injection, second dose of GnRH; and 16 h after second GnRH cows were TAI). Cows in the TAI group were resynchronized with an intravaginal insert containing progesterone inserted 18 d after TAI and removed 7 d later when GnRH was given. Cows were examined by ultrasonography on d 32 after TAI; nonpregnant cows received PGF_2α and GnRH 56 h later followed by TAI 16 h after the GnRH injection. Nonpregnant cows in TAI group were reinseminated up to 5 times using the same scheme. Cows in the NS group were exposed to bulls 14 d after the second PGF_2α injection, and ultrasonography was performed 42 d after exposure to bulls to determine pregnancy status. Nonpregnant cows in the NS group were reexamined by transrectal palpation combined with ultrasound every 28 d until diagnosed pregnant or 223 d postpartum, whichever occurred first. Cows diagnosed pregnant in TAI or NS were reconfirmed 28 d later to determine pregnancy loss. All bulls underwent an evaluation of breeding soundness and were rested for 14 d after 14 d of cow exposure. Health disorders were evaluated up to 70 d postpartum, and body condition score was evaluated at d 70 postpartum. Blood was collected on d 56 and 65 postpartum and analyzed for progesterone to determine cyclicity. The proportion of pregnant cows in the first 21 d of breeding did not differ between groups. The overall 21-d cycle pregnancy rate (PR), which included a total of 8 and 5 service opportunities for NS and TAI, respectively, was not different between groups (25.7 and 25.0% for NS and TAI, respectively). The daily rate of pregnancy was 15% greater for NS than TAI because cows in NS had a greater PR, which resulted in fewer median days open (111 vs. 116 d). Proportion of pregnant cows at 223 d postpartum was greater in the NS than TAI group (84.2 vs. 74.8%, respectively). Cyclicity did not affect reproductive performance. Cows with body condition score ≥2.75 had greater proportion of pregnant cows in the first 21 d of breeding and daily PR in the first 223 d postpartum Primiparous cows had greater proportion of pregnant cows and daily PR than multiparous cows at 223 d postpartum. In conclusion, the greater proportion of pregnant cows in the NS group was attributed to more opportunities for breeding than in the TAI group.     

Pregnancy in dairy cows after synchronized ovulation regimens with or without presynchronization and progesterone

Two experiments examined pregnancy after synchronized ovulation (Ovsynch) with or without progesterone (P4) administered via controlled internal drug release (CIDR) intravaginal inserts. In experiment 1, 262 lactating cows in one herd were in 3 treatments: Ovsynch (n = 91), Ovsynch + CIDR (n = 91), and control (n = 80). The Ovsynch protocol included injections of GnRH 7 d before and 48 h after an injection of PGF20. Timed artificial insemination (TAI; 57 to 77 d postpartum) was 16 to 20 h after the second GnRH injection. Cows in the Ovsynch + CIDR group also received a CIDR (1.9 g of P4) insert for 7 d starting at first GnRH injection. Control cows received A-I when estrus was detected using an electronic estrus detection system. Based on serum P4, 44.1% of cows were cyclic before Ovsynch. Pregnancy rates at 29 d (59.3 vs. 36.3%) and 57 d (45.1 vs. 19.8%) after TAI and embryo survival (75.9 vs. 54.5%) from 29 to 57 d were greater for Ovsynch + CIDR than for Ovsynch alone. In experiment 2, 630 cows in 2 herds received TAI at 59 to 79 d postpartum after 6 treatments. Estrous cycles were either presynchronized (2 injections of PGF2alpha 14 d apart; n = 318) or not presynchronized (n = 312). Within those groups, Ovsynch was initiated 12 d after second presynchronization PGF2alpha, and used alone (n = 318) or with CIDR inserts for 7 d (1.38 g of P4/insert, n = 124 or 1.9 g of P4/insert, n = 188). Before Ovsynch, 80% of cows were cyclic. Presynchronization increased pregnancy (46.8 vs. 37.5%) at 29 d after TAI, but CIDR inserts had no effect on pregnancy in experiment 2. Overall embryonic survival between 29 and 57 d in experiment 2 was 57.7%. Use of CIDR inserts with Ovsynch improved conception and embryo survival in experiment 1 but not in experiment 2, in part due to differing proportions of cyclic cows at the outset. Presynchronization before Ovsynch enhanced pregnancy rate.     

A resynchronization of ovulation program based on ovarian structures present at nonpregnancy diagnosis reduced time to pregnancy in lactating dairy cows

Our objective was to evaluate time to pregnancy after the first service postpartum and pregnancy per artificial insemination (P/AI) in dairy cows managed with 2 resynchronization of ovulation programs. After first service, lactating Holstein cows were blocked by parity (primiparous vs. multiparous) and randomly assigned to the d 32 Resynch (R32; n = 1,010) or short Resynch (SR; n = 1,000) treatments. Nonpregnancy diagnosis (NPD) was conducted 32 ± 3 d after AI by transrectal ultrasonography. Nonpregnant cows in R32 received the Ovsynch protocol: GnRH, PGF_2α 7 d later, GnRH 56 h later, and timed AI (TAI) 16 to 18 h later. Cows in SR with a corpus luteum (CL) ≥15 mm and a follicle ≥10 mm at NPD received PGF_2α, PGF_2α 24 h later, GnRH 32 h later, and TAI 16 to 18 h later. Cows in SR without a CL ≥15 mm or a follicle ≥10 mm at NPD received a modified Ovsynch protocol with 2 PGF_2α treatments and progesterone (P4) supplementation (GnRH plus CIDR, PGF_2α and CIDR removal 7 d later, PGF_2α 24 h later, GnRH 32 h later, and TAI 16 to 18 h later). Blood samples were collected from a subgroup of cows at the GnRH before TAI (R32 = 114; SR = 121) to measure P4 concentration. Binomial outcomes were analyzed with logistic regression and hazard of pregnancy (R32 = 485; SR = 462) with Cox's proportional regression in SAS (SAS Institute, Cary, NC). For P/AI analysis, the TAI service was the experimental unit (R32 = 720; SR = 819). Models included treatment and parity as fixed effects and farm as random effect. The hazard of pregnancy was greater for the SR treatment (hazard ratio = 1.18; 95% confidence interval: 1.01–1.37). Median time to pregnancy was 95 and 79 d for the R32 and SR treatments, respectively. At NPD, 71.3 and 71.2% of cows had a CL for the R32 and SR treatments, respectively. Treatment did not affect overall P/AI 32 ± 3 d after AI (R32 = 31.0% vs. SR = 33.9%) or for cows with a CL at NPD (R32 = 32.7% vs. SR = 32.8%). For cows with no CL at NPD, P/AI was greater for the SR treatment (36.9%) than for the R32 treatment (28.6%). Pregnancy loss from 32 to 63 d after AI was similar for all services combined (R32 = 8.3% vs. SR = 10.4%) and for cows with no CL at NPD (R32 = 13.2% vs. SR = 7.2%) but tended to be affected by treatment for cows with a CL at NPD (R32 = 6.8% vs. SR = 11.9%). Treatment affected the proportion of cows with P4 ≤0.5 ng/mL at the GnRH before TAI for all cows (R32 = 68.4% vs. SR = 81.8%), tended to have an effect among cows with a CL (R32 = 70.0% vs. SR = 81.8%), and had no effect for cows with no CL (R32 = 64.7% vs. SR = 81.8%). We concluded that the SR program reduced time to pregnancy because of a reduction of the interbreeding interval for cows with a CL at NPD and greater P/AI in cows with no CL at NPD.     

Effects of additional prostaglandin F2α and estradiol-17β during Ovsynch in lactating dairy cows

This study was designed to evaluate whether decreasing circulating progesterone (P4) or increasing circulating estradiol-17β (E2) near the time of artificial insemination (AI) in an Ovsynch protocol would increase pregnancies per AI (P/AI) in lactating dairy cows. Six hundred nineteen lactating Holstein cows (n = 772 inseminations) received Ovsynch (GnRH-7 d-PGF_2α-56 h-GnRH-16 h-timed AI). Cows were randomized in a 2 × 2 factorial experiment of 4 treatments to receive or not receive 25 mg of PGF_2α 24 h after the standard PGF_2α of Ovsynch, or 0.5 mg of E2 at the time of the final GnRH of Ovsynch, or both. Blood samples were collected 24 h after normal PGF_2α and at final GnRH to evaluate circulating P4. Ovarian ultrasound was done at final GnRH to determine preovulatory follicle size. Ovulation was confirmed by ultrasound 5 d after AI. Treatment with additional PGF_2α increased the percentage of cows that had complete luteal regression (95.6%) compared with control cows (84.6%). In contrast, additional PGF_2α had no detectable effect on P/AI (control = 41.5% vs. + PGF_2α = 44.7%). Supplementation with E2 increased expression of estrus (84.4 vs. 37.2%), but had no effect on overall fertility and even tended to have a negative effect on fertility in cows that ovulated to the second GnRH (control = 51.5% vs. +E2 = 44.0%). Thus, additional treatments with PGF_2α or E2 during Ovsynch can be used to increase synchronization and expression of estrus during Ovsynch, although the lack of improvement in fertility makes these treatments unwarranted.