Timed artificial insemination with estradiol cypionate or insemination at estrus in high-producing dairy cows

A total of 799 Holstein cows from 3 herds were randomly assigned at 37 +/- 3 d in milk (DIM) to timed artificial insemination (AI) or insemination at detected estrus. Cows were presynchronized with injections of PGF(2alpha) at 37 and 51 DIM. At 65 DIM, cows received an injection of GnRH, followed 7 d later by PGF(2alpha). Cows in the estrus-detected group were inseminated after being observed in estrus during the 7 d after the last PGF(2alpha). Cows in the timed AI group received an injection of 1 mg of estradiol cypionate (ECP) 24 h after the last PGF(2alpha). If detected in estrus or=1 ng/mL; L = <1 ng/mL), resulting in 8 combinations (LLL, LHL, LLH, LHH, HHH, HHL, HLH, and HLL). Conception rates and pregnancy rates were higher for cows in the timed AI group than in the estrus-detected group at 30, 44, and 58 d (e.g., at 58 d, pregnancy rates were 42.2% for multiparous cows or 34.4% for primiparous cows in the group receiving ECP and timed AI compared with only 20.8 or 18.8% for respective parity subgroups for the treatment group inseminated only at detected estrus). Pregnancy losses were 11.5% from 30 to 58 d and did not differ between treatments. Cyclic cows within both treatments had higher estrous responses, conception rates, and pregnancy rates. Cows that responded to presynchronization and to luteolysis (HHL) had the highest conception and pregnancy rates, followed by cows classified as LHL. Use of 1 mg of ECP to induce ovulation as part of a synchrony regimen improved reproduction at first postpartum insemination in dairy cows.     

Reproduction in dairy cows following progesterone insert presynchronization and resynchronization protocols

The objectives of this study were to evaluate the effects of an intravaginal insert containing progesterone (CIDR, controlled internal drug releasing) administered in presynchronization and resynchronization protocols on cyclicity, detection of estrus, pregnancy rate, and pregnancy loss to first AI; reinsemination patterns; and pregnancy rates to second postpartum AI before and after the time of first-service pregnancy diagnosis in dairy cows. Holstein cows (n = 1,052) were blocked by parity and BCS at 3 ± 3 d in milk (study d 0 = day of calving) and assigned randomly to 1 of 3 presynchronization treatments. During the presynchronization programs, all cows received 2 injections of PGF_2α, on study d 35 and 49. Cows enrolled in the control presynchronization treatment received AI after detected estrus from study d 49 to 62. Cows enrolled in the CIDR estrus-detection (CED) presynchronization treatment received a CIDR insert from study d 42 to 49 and AI on detection of estrus from d 49 to 62. Cows enrolled in the CIDR timed AI (CTAI) presynchronization treatment received the same treatment as CED, but were subjected to timed AI on study d 72 after the Ovsynch (GnRH, 7 d PGF_2α, 2 d GnRH, 24 h timed AI) protocol. The control and CED cows not inseminated by study d 62 were enrolled in the Ovsynch protocol on the same day and received timed AI on study d 72. After first AI, cows were assigned to no resynchronization (RCON) or resynchronization with a CIDR insert (RCIDR) between 14 and 21 d after AI. Blood samples collected on study d 35, 49, and 62 were analyzed for concentrations of progesterone and cows were classified as anestrous when progesterone was <1.0 ng/mL in the first 2 samples. On study d 62, anestrous cows with progesterone ≥ 1.0 ng/mL were classified as having resumed cyclicity. Pregnancy was diagnosed at 31 and 60 d after first AI and at 42 d after second AI. A greater proportion of anestrous cows in CED and CTAI became cyclic by d 62 postpartum than control cows. Resynchronization with the CIDR insert increased the pregnancy rate at 31 d after first AI in CED and CTAI, and at 60 d after AI in all cows because of reduced pregnancy loss. These results indicate that presynchronization with the CIDR insert increased induction of cyclicity in anestrous cows and that resynchronization with the CIDR insert did not affect the reinsemination rate but did reduce pregnancy loss and increased the pregnancy rate at 60 d after first AI.     

Effect of early or late resynchronization based on different methods of pregnancy diagnosis on reproductive performance of dairy cows

The aim of this study was to compare the reproductive performance of dairy cows subjected to early (ER) or late (LR) resynchronization programs after nonpregnancy diagnoses based on either pregnancy-associated glycoproteins (PAG) ELISA or transrectal palpation, respectively. In addition, the accuracy of the PAG ELISA for early pregnancy diagnosis was assessed. Lactating Holstein cows were subjected to a Presynch-Ovsynch protocol with timed artificial insemination (AI) performed between 61 and 74 DIM. On the day of the first postpartum AI, 1,093 cows were blocked by parity and assigned randomly to treatments; however, because of attrition, 452 ER and 520 LR cows were considered for the statistical analyses. After the first postpartum AI, cows were observed daily for signs of estrus and inseminated on the same day of detected estrus. Cows from ER that were not reinseminated in estrus received the first GnRH injection of the Ovsynch protocol for resynchronization 2 d before pregnancy diagnosis. On d 28 after the previous AI (d 27 to 34), pregnancy status was determined by PAG ELISA, and nonpregnant cows continued on the Ovsynch protocol for reinsemination. Pregnant cows had pregnancy status reconfirmed on d 46 after AI (d 35 to 52) by transrectal palpation, and those that lost the pregnancies were resynchronized. Cows assigned to LR had pregnancy diagnosed by transrectal palpation on d 46 after AI (d 35 to 52) and nonpregnant cows were resynchronized with the Ovsynch protocol. Blood was sampled on d 28 after AI (d 27 to 34) from cows in both treatments that had not been reinseminated on estrus and again on d 46 after AI (d 35 to 52) for assessment of PAG ELISA to determine the accuracy of the test. Cows were subjected to treatments for 72 d after the first insemination. Pregnancy per AI (P/AI) at first postpartum timed AI did not differ between treatments and averaged 28.9%. The proportion of nonpregnant cows that were resynchronized and received timed AI was greater for ER than for LR (30.0 vs. 7.6%). Cows in ER had a shorter interval between inseminations when inseminated following spontaneous estrus (21.7 ± 1.1 vs. 27.8 ± 0.8 d) or after timed AI (35.3 ± 1.2 vs. 55.2 ± 1.4 d). Nevertheless, the ER did not affect the rate of pregnancy (adjusted hazard ratio = 1.23; 95% confidence interval = 0.94 to 1.61) or the median days postpartum to pregnancy (ER = 132 vs. LR = 140). A total of 2,129 PAG ELISA were evaluated. Overall, sensitivity, specificity, and positive and negative predictive values averaged 95.1, 89.0, 90.1, and 94.5%, respectively, and the accuracy was 92.1%. In conclusion, PAG ELISA for early diagnosis of pregnancy had acceptable accuracy, but early resynchronization after nonpregnancy diagnosis with PAG ELISA did not improve the rate of pregnancy or reduce days open in dairy cows continuously observed for estrus.     

Fertility of dairy cows after resynchronization of ovulation at three intervals following first timed insemination

Lactating Holstein cows (n = 711) on a commercial dairy farm in Wisconsin received a hormonal synchronization protocol to initiate first timed artificial insemination (TAI) on the following postpartum schedule: two injections of 25 mg PGF2alpha at 32 +/- 3 d and 46 +/- 3 d (Presynch); 100 microg GnRH at 60 +/- 3 d; 25 mg PGF2alpha at 67 +/- 3 d; and 100 microg GnRH + TAI at 69 +/- 3 d (Ovsynch). At first TAI, cows were randomly assigned to initiate the first GnRH injection of a hormonal protocol for resynchronization of ovulation (Resynch; 100 microg GnRH, d 0, 25 mg PGF2alpha, d 7, 100 microg GnRH + TAI, d 9) at 19 (D19), 26 (D26), or 33 d (D33) after first TAI to set up a second TAI service for cows failing to conceive to Ovsynch. Overall pregnancy rate per artificial insemination (PR/AI) to Ovsynch assessed 68 d after TAI was 31% and did not differ among treatment groups. For Resynch, PR/AI was assessed 26 d after TAI for D19 and D26 cows and 33 d after TAI for D33 cows. Overall PR/AI to Resynch was 32%. However, the PR/AI for D26 (34%) and D33 (38%) cows to Resynch was greater than for D19 cows (23%). Cows with a CL at the PGF2alpha injection (D19 cows) or at the first GnRH injection (D26 + D33 cows) of Resynch exhibited greater PR/AI to Resynch compared with cows without a CL. Survival analysis (failure time) of cows in the D26 and D33 treatment groups across the first three TAI services did not differ statistically. Although administration of GnRH to pregnant cows 19 d after first TAI service did not appear to induce iatrogenic embryonic loss, initiation of Resynch 19 d after first TAI service resulted in a lower PR/AI compared with initiation of Resynch 26 or 33 d after first TAI service.     

Supplemental progesterone and timing of resynchronization on pregnancy outcomes in lactating dairy cows

The objective was to determine the effect of exogenous progesterone (P₄) in a timed artificial insemination (TAI) protocol initiated at 2 different times post-AI on pregnancies per AI (P/AI) in lactating dairy cows. Cows (n = 1,982) in 5 dairy herds were assigned randomly at a nonpregnancy diagnosis 32 ± 3 d post-AI to 1 of 4 resynchronization (RES) treatments arranged in a 2 × 2 factorial design using the Ovsynch-56 (GnRH, 7 d later PGF_2α, 56 h later GnRH, 16 h later TAI) protocol. Treatments were as follows: cows initiating RES 32 ± 3 d after AI with no supplemental P₄ (d 32 RES-CON; n = 516); same as d 32 RES-CON plus a controlled internal drug release (CIDR) insert containing P₄ at the onset of Ovsynch-56 (d 32 RES-CIDR; n = 503); cows initiating RES 39 ± 3 d after AI (d 39 RES-CON; n = 494); and same as d 39 RES-CON plus a CIDR (d 39 RES-CIDR; n = 491). Cows were inseminated if observed in estrus before TAI. The P/AI was determined 32 and 60 d after TAI. In a subgroup of cows (n = 1,152), blood samples were collected and ovarian structures examined by ultrasonography on the days of the first GnRH (G1) and PGF_2α of Ovsynch-56. Percentage of cows with a corpus luteum (CL) at G1 was unaffected by timing of treatments, but percentage of cows with a CL at PGF_2α was greater for d 32 than for d 39 cows (87.9 vs. 79.4%). In addition, percentage of cows with P₄ ≥1 ng/mL at G1 was unaffected by timing of treatments, but was increased for d 32 compared with d 39 RES cows on the day of the PGF_2α of the RES protocols (86.5 vs. 74.3%). Treatment did not affect ovulation to G1 or P/AI 32 d after RES TAI (d 32 RES-CON = 30.1%, d 32 RES-CIDR = 28.8%, d 39 RES-CON = 27.5%, d 39 RES-CIDR = 30.5%). A greater percentage of d 39 RES cows underwent premature luteolysis during the RES protocol compared with d 32 RES cows. An interaction was detected between day of RES initiation and CIDR treatment, in which the CIDR increased P/AI 60 d after TAI for d 39 (CON = 23.7% vs. CIDR = 28.0%), but not for d 32 (CON = 26.9% and CIDR = 24.2%) cows. Pregnancy loss was unaffected by treatment. In addition, cows had improved P/AI 60 d after TAI when they received a CIDR and did not have a CL (CON-CL = 28.2%, CON-No CL = 19.2%, CIDR-CL = 27.0%, and CIDR-No CL = 26.5%) or had P₄ <1 ng/mL (CON-High P₄ = 27.8%, CON-Low P₄ = 15.0%, CIDR-High P₄ = 25.0%, and CIDR-Low P₄ = 29.4%) at G1, but not if a CL was present or P₄ was ≥1 ng/mL at G1. In conclusion, addition of a CIDR insert to supplement P₄ during the RES protocol increased P/AI for cows initiating RES 39 ± 3 d after AI but not 32 ± 3 d after AI.     

Resynchronizing estrus with progesterone or progesterone plus estrogen in cows of unknown pregnancy status

Two experiments were conducted to test 2 progesterone (P4)-based treatments that were applied to lactating dairy cattle of unknown pregnancy status to resynchronize estrus of nonpregnant cows. In experiment 1, cows were assigned randomly before a timed AI (TAI) to 1) treatment with a CIDR (controlled internal drug-releasing intravaginal insert containing P4) for 7 d starting on d 13 after TAI (CIDR; n = 300) or 2) no P4 treatment (control; n = 330). Compared with controls, P4 increased the synchrony of those detected in estrus, but failed to increase the overall return rates of non-pregnant cows during the 6 d after CIDR removal (27% vs. 31%; d 20 to 26 after TAI) and did not alter synchronized conception rates (32% vs. 20%) of those inseminated. Use of P4 did not compromise pregnancies resulting from TAI compared with controls (38% vs. 42%), but increased embryo survival between d 29 and 57 after TAI (65.5% vs. 44.3%). In experiment 2, on d 13 after TAI, 196 cows were treated with a CIDR insert for 7 d. Controls received no further treatment. Remaining cows were treated with 1 of 3 estrogen regimens: 1 mg of estradiol benzoate (EB), 0.5 mg of estradiol cypionate (ECP), or 1 mg of ECP on both d 13 and 21. Only 60% of nonpregnant, estrogen-treated cows were detected in estrus between d 20 and 26, and rates of return and conception did not differ among treatments. Estrogen on d 13 did not consistently turn over the dominant follicle when given at CIDR insertion but did increase concentrations of estradiol and reduced luteal function when administered on d 13 and 21 (24 h after CIDR removal). Treatments had no negative effects on milk yield, dry matter intake, or established pregnancies. Use of P4 alone had little effect on overall rates of return to estrus or conception at the first eligible estrus in experiment 1. Combining estrogen with P4 in experiment 2 had no detrimental effects on established pregnancies or subsequent conception and failed to improve return rates beyond P4 alone.     

Effects of resynchronization programs on pregnancy per artificial insemination, progesterone, and pregnancy-associated glycoproteins in plasma of lactating dairy cows

Objectives were to develop a timed artificial insemination (TAI) resynchronization program to improve pregnancy per AI and to evaluate responses of circulating progesterone and pregnancy-associated glycoproteins in lactating cows. Cows (n = 1,578) were presynchronized with 2 injections of PGF_2α, given 14 d apart starting on d 45 ± 3 postpartum, followed by Ovsynch [2 injections of GnRH 7 d before and 56 h after injection of PGF_2α, TAI 16 h after second injection (d 0)]. The Resynch-treated cows received an intravaginal progesterone insert from d 18 to 25, GnRH on d 25, and pregnancy diagnosis on d 32, and nonpregnant cows received PGF_2α., GnRH 56 h later, and TAI 16 h later (d 35). The control cows were diagnosed for pregnancy on d 32 and nonpregnant cows received GnRH, PGF_2α 39 d after TAI, GnRH 56 h later, and TAI 16 h later (d 42). Pregnancy was reconfirmed on d 60 after AI. Ovarian structures were examined in a subset of cows at the time of GnRH and PGF_2α injections. Blood samples for analyses of progesterone and pregnancy-associated glycoproteins were collected every 2 d from d 18 to 30 in 100 cows, and collection continued weekly to d 60 for pregnant cows (n = 43). Preenrollment pregnancies per AI on d 32 did not differ for cows subsequently treated as Resynch (45.8%, n = 814) and control (45.9%, n = 764), and pregnancy losses on d 60 were 6.7 and 4.0%, respectively. Resynchronized service pregnancy per AI (36%, n = 441; 39.5%, n = 412) and pregnancy losses (6.3 and 6.7%) did not differ for Resynch and control treatments, respectively. Days open for pregnant cows after 2 TAI were less for the Resynch treatment than for the control treatment (96.2 ± 0.82 vs. 99.5 ± 0.83 d). Cows in the Resynch treatment had more large follicles at the time of GnRH. The number of corpora lutea did not differ between treatments at the time of PGF_2α. Plasma progesterone for pregnant cows was greater for Resynch cows than for control cows (18-60 d; 6.6 vs. 5.3 ng/mL), and plasma concentrations of progesterone on d 18 were greater for pregnant cows than for nonpregnant cows (5.3 vs. 4.3 ng/mL). Plasma pregnancy-associated glycoproteins during pregnancy were lower for cows in the Resynch treatment compared with control cows on d 39 (2.8 vs. 4.1 ng/mL) and 46 (1.3 vs. 3.0 ng/mL). Cows pregnant on d 32 that lost pregnancy by d 60 (n = 7) had lower plasma concentrations of pregnancy-associated glycoproteins on d 30 than cows that maintained pregnancy (n = 36; 2.9 vs. 5.0 ng/mL). Pregnancy-associated glycoproteins on d 30 (>0.33 ng/mL) were predictive of a positive d 32 pregnancy diagnosis (sensitivity = 100%; specificity = 90.6%). In conclusion, Resynch and control protocols had comparable pregnancy per AI for first and second TAI services, but pregnancy occurred 3.2 d earlier in the Resynch group because inseminations in the Resynch treatment began 7 d before those in the control treatment. Administration of an intravaginal progesterone insert, or GnRH, or both increased progesterone during pregnancy. Dynamics of pregnancy-associated glycoproteins were indicative of pregnancy status and pregnancy loss.     

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.     

Resynchronization strategies to improve fertility in lactating dairy cows utilizing a presynchronization injection of GnRH or supplemental progesterone: I. Pregnancy rates and ovarian responses

Objectives were to evaluate 3 resynchronization protocols for lactating dairy cows. At 32 ± 3 d after pre-enrollment artificial insemination (AI; study d −7), 1 wk before pregnancy diagnosis, cows from 2 farms were enrolled and randomly assigned to 1 of 3 resynchronization protocols after balancing for parity, days in milk, and number of previous AI. All cows were examined for pregnancy at 39 ± 3 d after pre-enrollment AI (study d 0). Cows enrolled as controls (n = 386) diagnosed not pregnant were submitted to a resynchronization protocol (d 0-GnRH, d 7-PGF_2α, and d 10-GnRH and AI) on the same day. Cows enrolled in the GGPG (GnRH-GnRH-PGF_2α-GnRH) treatment (n = 357) received a GnRH injection at enrollment (d −7) and if diagnosed not pregnant were submitted to the resynchronization protocol for control cows on d 0. Cows enrolled in CIDR treatment (n = 316) diagnosed not pregnant received the resynchronization protocol described for control cows with addition of a controlled internal drug release (CIDR) insert containing progesterone (P₄) from d 0 to 7. In a subgroup of cows, ovaries were scanned and blood was sampled for P₄ concentration on d 0 and 7. After resynchronized AI, cows were diagnosed for pregnancy at 39 ± 3 and 67 ± 3 d (California herds) or 120 ± 3 d (Arizona herds). Cows in the GGPG treatment had more corpora lutea than CIDR and control cows on d 0 (1.30 ± 0.11, 1.05 ± 0.11, and 1.05 ± 0.11, respectively) and d 7 (1.41 ± 0.14, 0.97 ± 0.13, and 1.03 ± 0.14, respectively). A greater percentage of GGPG cows ovulated to GnRH given on d 0 compared with CIDR and control cows (48.4, 29.6, and 36.6%, respectively), but CIDR and control did not differ. At 39 ± 3 d after resynchronized AI, pregnancy per AI (P/AI) was increased in GGPG (33.6%) and CIDR (31.3%) cows compared with control (24.6%) cows. At 67 or 120 ± 3 d after resynchronized AI, P/AI of GGPG and CIDR cows was increased compared with control cows (31.2, 29.5, and 22.1%, respectively). Presynchronizing the estrous cycle of lactating dairy cows with a GnRH 7 d before the start of the resynchronization protocol or use of a CIDR insert within the resynchronization protocol resulted in greater P/AI after resynchronized AI compared with control cows.     

Pregnancy outcomes are not improved by administering gonadotropin-releasing hormone at initiation of a 5-day CIDR-Cosynch resynchronization protocol for lactating dairy cows

Using a 5-d controlled internal drug-release (CIDR)-Cosynch resynchronization protocol, the objective of this study was to determine the effect of the initial GnRH injection on pregnancy per artificial insemination (P/AI) to the second artificial insemination in lactating Holstein dairy cows. On 37 ± 3 d (mean ± standard deviation) after the first artificial insemination, and upon nonpregnancy diagnosis (d 0 of the experiment), lactating cows eligible for a second artificial insemination (n = 429) were enrolled in a 5-d CIDR-Cosynch protocol. On d 0, all cows received a CIDR insert and were assigned randomly to receive the initial GnRH injection (GnRH; n = 226) of the protocol or no-GnRH (n = 203). Blood samples were collected from a sub-group of cows (n = 184) on d 0 and analyzed for progesterone (P4) concentration. On d 5, CIDR inserts were removed, and all cows received 1 injection of PGF_2α. On d 6 and 7, cows were observed once daily by employees for tail-chalk removal, and cows detected in estrus on d 6 or 7 received artificial insemination that day (EDAI), and did not receive the final GnRH injection. The remaining cows not detected in estrus by d 8 received GnRH and timed artificial insemination (TAI). Pregnancy status was confirmed by transrectal palpation of uterine contents at 37 ± 3 d (mean ± standard deviation) after the second artificial insemination. Eliminating the initial GnRH injection had no effect on P/AI compared with cows receiving GnRH (27 vs. 21%), respectively. Similarly, method of insemination (EDAI vs. TAI) and its interaction with treatment had no effect on P/AI. Primiparous cows had greater P/AI than multiparous cows (31 vs. 21%). Mean P4 concentrations (n = 184) at the initiation of the protocol did not differ between treatments (4.51 ± 0.35 ng/mL no-GnRH vs. 3.96 ± 0.34 ng/mL of GnRH). When P4 concentrations were categorized as high (≥1 ng/mL) or low (<1 ng/mL), P/AI tended to be greater for high P4 concentrations (n = 136) compared with low (n = 48) P4 concentrations (26 vs. 16%, respectively). No differences were observed in the proportion of cows with high or low P4 between treatments. Collectively, these results provide evidence that eliminating the initial GnRH in a 5-d CIDR-Cosynch resynchronization protocol for lactating dairy cows did not reduce P/AI in this study.     

Effects of supplemental progesterone after artificial insemination on expression of interferon-stimulated genes and fertility in dairy cows

The objectives of the current study were to evaluate the effects of supplemental progesterone after artificial insemination (AI) on expression of IFN-stimulated genes (ISG) in blood leukocytes and fertility in lactating dairy cows. Weekly cohorts of Holstein cows were blocked by parity (575 primiparous and 923 multiparous) and method of insemination (timed AI or AI on estrus) and allocated randomly within each block to untreated controls, a controlled internal drug release (CIDR) containing 1.38 g of progesterone from d 4 to 18 after AI (CIDR4), or a CIDR on d 4 and another on d 7 after AI and both removed on d 18 (CIDR4+7). Blood was sampled to quantify progesterone concentrations in plasma and mRNA expression in leukocytes for the ubiquitin-like IFN-stimulated gene 15-kDa protein (ISG15) and receptor transporter protein-4 (RTP4) genes. Pregnancy was diagnosed on d 34 ± 3 and 62 ± 3 after AI. Treatment increased progesterone concentrations between d 5 and 18 after AI in a dose-dependent manner (control = 3.42, CIDR4 = 4.97, and CIDR4+7 = 5.46 ng/mL). Cows supplemented with progesterone tended to have increased luteolysis by d 19 after AI (control = 17.2; CIDR4 = 29.1; CIDR4+7 = 30.2%), which resulted in a shorter AI interval for those reinseminated after study d 18. Pregnancy upregulated expression of ISG in leukocytes on d 19 of gestation, but supplementing progesterone did not increase mRNA abundance for ISG15 and RTP4 on d 16 after insemination and tended to reduce mRNA expression on d 19 after AI. For RTP4 on d 19, the negative effect of supplemental progesterone was observed only in the nonpregnant cows. No overall effect of treatment was observed on pregnancy per AI on d 62 after insemination and averaged 28.6, 32.7, and 29.5% for control, CIDR4, and CIDR4+7, respectively. Interestingly, an interaction between level of supplemental progesterone and method of AI was observed for pregnancy per AI. For cows receiving exogenous progesterone, the lower supplementation with CIDR4 increased pregnancy per AI on d 62 in cows inseminated following timed AI (CIDR4 = 39.2; CIDR4+7 = 27.5%); in those inseminated following detection of estrus, however, the use of a second insert on d 7 resulted in greater pregnancy per AI (CIDR4 = 26.9; CIDR4+7 = 31.5%). Pregnancy loss did not differ among treatments. Supplemental progesterone post-AI using a single intravaginal insert on d 4 was beneficial to pregnancy in cows inseminated following timed AI, but incremental progesterone with a second insert on d 7 did not improve fertility of dairy cows.     

Intravaginal progesterone insert to synchronize return to estrus of previously inseminated dairy cows

An intravaginal progesterone insert (CIDR insert; 1.38 g of progesterone) was evaluated for synchronization of returns to estrus (SR), conception rate (CR), and pregnancy rate (PR) in dairy cows previously artificially inseminated (AI). Healthy, nonpregnant, lactating Holstein cows, > or = 40 and < or = 150 d postpartum at eight commercial farms were used. Cows detected in estrus and receiving AI 2, 3, or 4 d after one injection of PGF2alpha (25 mg) were assigned as either controls (n = 945), or to receive a CIDR insert (n = 948) for 7 d (14 to 21 +/- 1 d after AI). Cows were observed for returns to estrus from 18 to 26 +/- 1 d after initial AI (resynchrony period) and were reinseminated if in estrus. Vaginal mucus on CIDR inserts (97.3% retention) at removal was scored: 1 = no mucus; 2 = clear; 3 = cloudy; 4 = yellow; and 5 = red or brown. Percentage of cows in estrus (SR) during the 3 d after CIDR insert removal was contrasted to the highest 3-d cumulative percentage in estrus for controls. Cows conceiving to initial AI were omitted in calculations of SR, CR, and PR during resynchrony. Mucous scores of 3 or 4 (mild irritation) were observed in 65% of cows and a score of 5 (more severe irritation) was observed in 2%; otherwise, health was unaffected. The PR to initial AI was lower for cows subsequently receiving CIDR inserts than for controls (32.7 vs. 36.7%). The CIDR insert increased SR (34.1 vs. 19.3% in 3 d) and overall estrus detection (43% in 4 d vs. 36% in 9 d) compared with controls. For the 9-d resynchrony period, CR and PR for CIDR-treated (26.7, 12.2%) and control (30.9, 11.1%) cows did not differ significantly. The CIDR inserts improved synchrony of returns to estrus, slightly reduced PR to initial AI, but did not affect CR or PR to AI during the resynchrony period.     

Effect of time of artificial insemination and supplemental estradiol on reproduction of lactating dairy cows

Our objectives were to compare reproductive responses of dairy cows receiving timed artificial insemination (AI) either at 48 or 72 h after induction of luteolysis and supplemented or not with estradiol cypionate (ECP). Holstein cows (971) had their estrous cycles presynchronized with injections of PGF_2α at 37 and 51 d in milk (DIM) and then received an injection of GnRH at 64 DIM and an injection of PGF_2α at 71 DIM. Cows were then assigned to a 2 × 2 factorial randomized block experiment; cows in the CoSynch 48 h (CoS48) received a final injection of GnRH concurrent with timed AI 48 h after PGF_2α, whereas cows in the CoSynch 72 h (CoS72) received GnRH and timed AI 72 h after PGF_2α. Half of the cows in each CoSynch protocol received an injection of 1 mg of ECP 24 h after PGF_2α. Therefore, the 4 treatments were as follows: CoS48-NECP (n = 240), CoS72-NECP (n = 246), CoS48-ECP (n = 245), and CoS72-ECP (n = 240). Blood was sampled at 7 d before and at the first GnRH of the CoSynch from all cows for analysis of progesterone concentration in plasma. Cows were classified as anovular when progesterone was less than 1.0 ng/mL in both samples. Blood was also sampled during proestrus from a subset of 123 cows to measure concentrations of estradiol and at 7 d after timed AI to measure concentrations of progesterone. Ovaries from the same subset of 123 cows were examined by ultrasonography to determine ovulatory follicle diameter and incidence of ovulation. Pregnancy was diagnosed at 40 and 68 d after AI. Prevalence of cyclic cows was 72.4% and was similar among treatments. Concentrations of estradiol increased after ECP treatment and at 72 h of proestrus with CoS72. Pregnancy at 40 and 68 d after AI and pregnancy loss were not affected by timing of AI or supplemental ECP. Delaying timed AI to 72 h and supplementation with ECP increased the proportion of cows displaying estrus at AI, and cows detected in estrus had increased pregnancy per AI associated with improved ovulation and increased postovulatory progesterone concentration. These results indicate that extending the proestrus by delaying timed AI from 48 to 72 h plus supplemental ECP, despite increased expression of estrus at timed AI, did not improve reproductive performance of lactating dairy cows at first AI.     

Effect of interval between induction of ovulation and artificial insemination (AI) and supplemental progesterone for resynchronization on fertility of dairy cows subjected to a 5-d timed AI program

Objectives were to investigate 2 intervals from induction of ovulation to artificial insemination (AI) and the effect of supplemental progesterone for resynchronization on fertility of lactating dairy cows subjected to a 5-d timed AI program. In experiment 1, 1,227 Holstein cows had their estrous cycles presynchronized with 2 injections of PGF_2α at 46 and 60 d in milk (DIM). The timed AI protocols were initiated with GnRH at 72 DIM, followed by 2 injections of PGF_2α at 77 and 78 DIM and a second injection of GnRH at either 56 (OVS56) or 72 h (COS72) after the first PGF_2α of the timed AI protocols. All cows were time-inseminated at 72 h after the first PGF_2α injection. Pregnancy was diagnosed on d 32 and 60 after AI. In experiment 2, 675 nonpregnant Holstein cows had their estrous cycles resynchronized starting at 34 d after the first AI. Cows received the OVS56 with (RCIDR) or without (RCON) supplemental progesterone, as an intravaginal insert, from the first GnRH to the first PGF_2α. Pregnancy diagnoses were performed on d 32 and 60 after AI. During experiment 2, subsets of cows had their ovaries scanned by ultrasonography at the first GnRH, the first PGF_2α, and second GnRH injections of the protocol. Blood was sampled on the day of AI and 7 d later, and concentrations of progesterone were determined in plasma. Cows were considered to have a synchronized ovulation if they had progesterone <1 and >2.26 ng/mL on the day of AI and 7 d later, respectively, and if no ovulation was detected between the first PGF_2α and second GnRH injections during resynchronization. In experiment 1, the proportion of cows detected in estrus at AI was greater for COS72 than OVS56 (40.6 vs. 32.4%). Pregnancy per AI (P/AI) did not differ between OVS56 (46.4%) and COS72 (45.5%). In experiment 2, cows supplemented with progesterone had greater P/AI compared with unsupplemented cows (51.3 vs. 43.1%). Premature ovulation tended to be greater for RCON than RCIDR cows (7.5 vs. 3.6%), although synchronization of the estrous cycle after timed AI was similar between treatments. Timing of induction of ovulation with GnRH relative to insemination did not affect P/AI of dairy cows enrolled in a 5-d timed AI program. Furthermore, during resynchronization starting on d 34 after the first AI, supplementation with progesterone improved P/AI in cows subjected to the 5-d timed AI protocol.     

Hyperketolactia occurrence before or after artificial insemination is associated with a decreased pregnancy per artificial insemination in dairy cows

The reproductive parameters of dairy cattle have continuously declined worldwide over the last 50 years. Nutritional imbalances are identified as risk factors for this decrease of reproductive performance. The present paper aims to quantify the decrease in the pregnancy per artificial insemination (P/AI) in the case of high milk ketones before and after AI. A total of 388,731 test-day from the Brittany Milk Recording Program in France from 226,429 cow-lactations were provided for this trial. For each test-day, information about lactation characteristics, date of AI, date of the following calving, and acetone and β-hydroxybutyrate (BHB) values were included. Ketones were predicted by Fourier transform mid-infrared spectroscopy using MilkoScan Foss analyzers (Foss, Hillerød, Denmark). Many thresholds were evaluated to define cows with hyperketolactia. Hyperketolactia statuses were then categorized into 1 of 4 possible classes according to the milk ketone dynamics for each AI and each threshold of acetone or BHB values (low-low, high-low, low-high, and high-high) within 20 d before and after AI. Similarly, the dynamics of udder health were characterized by changes in somatic cell counts measured at the same test day as ketone bodies. A logistic regression with a Poisson correction was performed to explain the relationship of P/AI with milk ketones and somatic cell count dynamics. Predicted acetone and BHB ranged from −0.51 to 4.92 mM (mean = 0.08 mM, SD = 0.10 mM) and −0.62 to 5.85 mM (mean = 0.07 mM, SD = 0.1 mM), respectively. Hyperketolactia defined by high acetone levels before AI was not associated with decreased P/AI, but high acetone levels after AI were associated with a >10% reduction in P/AI for all thresholds >0.10 mM. Hyperketolactia, defined by high BHB values before, after, or before and after AI, was associated with a 6 to 14% reduction in P/AI compared with cows with low BHB values. These associations are lower than those reported in previous trials in which blood ketones were used. High ketones in advanced lactation are likely to be the result of various primary disorders (secondary ketosis). Because the present work demonstrated that this situation is considered a risk factor for deteriorated reproductive performance, we suggest that high ketones in early and advanced lactation should be of interest to farm advisors.     

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.     

Effect of manipulating progesterone before timed artificial insemination on reproductive and endocrine outcomes in high-producing multiparous Holstein cows

Our objective was to evaluate the effect of manipulating progesterone (P4) concentrations before timed artificial insemination (TAI) on reproductive and endocrine outcomes in high-producing Holstein cows. Multiparous lactating Holstein cows (n = 80) were synchronized for first TAI using a Double-Ovsynch protocol and were randomly assigned to receive 25 mg of PGF_2α 1 d after the first GnRH treatment of the Breeding-Ovsynch protocol that included a once-used P4 insert (low-P4 group) or to receive 2 new P4 inserts during the Breeding-Ovsynch protocol (high-P4 group). Blood samples were collected thrice weekly from ?10 to 32 d relative to TAI for all cows and from 32 to 67 d after TAI for pregnant cows and were analyzed for P4 and pregnancy-specific protein B (PSPB) concentrations. Expression of IFNτ-stimulated gene 15 (ISG15) was assessed in blood leukocytes 18 and 20 d after TAI. As expected, P4 concentrations were greater for high-P4 cows than for low-P4 cows from 3 to 8 d before TAI. Incidence of double ovulation was 3-fold greater for low-P4 cows than for high-P4 cows (33 vs. 10%), which resulted in more twin pregnancies 32 d after TAI for low-P4 cows than for high-P4 cows (29 vs. 0%). Low-P4 cows had larger preovulatory follicles at the last GnRH treatment of the Double-Ovsynch protocol and greater P4 concentrations than high-P4 cows after TAI. Relative expression of ISG15 mRNA 18 and 20 d after TAI was greater for low-P4 cows than for high-P4 cows and for pregnant cows than for nonpregnant cows. Overall, PSPB concentrations tended to be greater for low-P4 cows than for high-P4 cows, and pregnant cows had greater P4 concentrations than nonpregnant cows. In summary, cows with low P4 before TAI had increased preovulatory follicle diameter, PSPB concentrations, relative expression of ISG15 mRNA 18 and 20 d after TAI, double ovulations, and twinning compared with cows with high P4 before TAI. Increasing P4 before TAI may effectively decrease double ovulation and twinning in high-producing multiparous Holstein cows.     

Ovarian structures and circulating steroids in heifers and lactating cows in summer and lactating and dry cows in winter

Two experiments compared follicular and luteal development and circulating steroid concentrations from induced luteolysis to ovulation in lactating Holstein cows (n = 27; 40.0 +/- 1.5 kg milk/day) vs. nulliparous heifers (n = 28; 11 to 17 mo-old) during summer (Experiment 1), and in lactating (n = 27; 45.9 +/- 1.4 kg milk/d) vs. dry cows (n = 26) during winter (experiment 2). All females received PGF2,, 6 d after ovulation and were monitored until next ovulation by daily ultrasound and assay of serum progesterone (P4) and estradiol (E2). Every female was used two or three times. In Experiment 1, lactating cows had high incidence of multiple ovulation (63.5%) compared with heifers (1.3%). Among single ovulators, there was no difference in maximal size of ovulatory follicles between lactating cows and heifers (15.8 vs. 16.5 mm, respectively). However, lactating cows had lower peak serum E2 (8.6 vs. 12.1 pg/ml), took longer to ovulate after luteolysis (4.6 vs. 3.8 d), developed more luteal tissue volume (7,293.6 vs. 5,515.2 mm3), and had lower serum P4 on d 6 after ovulation (2.0 vs. 3.0 ng/ml) than heifers (data included multiple ovulators). In experiment 2, multiple ovulations were similar between lactating and dry cows (17.9 vs. 17.2%, respectively). Peak serum E2 was also similar between lactating and dry cows (7.6 vs. 8.5 pg/ml) although lactating cows had larger ovulatory follicles (18.6 vs. 16.2 +/- 0.4 mm). Lactating cows took longer to ovulate (4.8 vs. 4.2 d), developed more luteal tissue (7,599 vs. 5,139 +/- 468 mm3), but had similar serum P4 (2.2 vs. 1.9 ng/ ml) compared with dry cows. Therefore, lactating cows had similar or lower circulating steroid concentrations than dry cows or heifers, respectively, despite having larger ovarian structures.     

Effect of interval from timed artificial insemination to initiation of resynchronization of ovulation on fertility of lactating dairy cows

To compare 2 strategies for systematically resynchronizing ovulation, lactating Holstein cows (n = 763) at various days in milk and prior artificial insemination services were assigned randomly at timed AI (TAI) to receive the first GnRH injection of Ovsynch 26 (D26) or 33 (D33) d after TAI to resynchronize ovulation (Resynch) in cows failing to conceive. Cows in the D26 treatment received GnRH 26 d after TAI and continued Resynch only when diagnosed not pregnant by using ultrasonography 33 d after TAI, whereas D33 cows initiated Resynch only when diagnosed not pregnant 33 d after TAI. Cows were classified based on the presence or absence of a corpus luteum (CL) at the not-pregnant diagnosis, and cows without a CL received an intravaginal progesterone-releasing insert during Resynch. When analyzed as a systematic strategy, pregnancy rate per AI (PR/AI) was greater for cows assigned to the D33 than the D26 Resynch treatment (39.4 vs. 28.6%). A treatment × parity interaction was detected for PR/AI after Resynch for nonpregnant cows having a CL in which primiparous cows had a greater PR/AI than multiparous cows when Resynch was initiated 33 d after the initial TAI, and primiparous and multiparous cows when Resynch was initiated 26 d after the initial TAI. Pregnancy loss for Resynch was 6.4% between 33 and 40 d, and 2.6% between 40 and 61 d after Resynch TAI. We concluded that delaying initiation of Resynch until 33 d after TAI increased PR/AI for primiparous cows.     

Atypical progesterone profiles and fertility in Swedish dairy cows

The incidence of normal and atypical progesterone profiles in Swedish dairy cows was studied. Data were collected from an experimental herd over 15 yr, and included 1,049 postpartum periods from 183 Swedish Holstein and 326 Swedish Red and White dairy cows. Milk progesterone samples were taken twice weekly until initiation of cyclical ovarian activity and less frequently thereafter. Progesterone profiles were 1) normal profile: first rise in milk progesterone above the threshold value before d 56 postpartum, followed by regular cyclical ovarian activity (70.4%); 2) delayed onset of cyclical ovarian activity: low milk progesterone the first 56 d postpartum (15.6%); 3) cessation of cyclical ovarian activity: ovarian activity resumed within 56 d postpartum, but ceased for a period of 14 d or more (6.6%); and 4) prolonged luteal phase: ovarian activity resumed within 56 d postpartum, but milk progesterone remained elevated in the nonpregnant cow for a period of 20 d or more (7.3%). Swedish Holsteins had 1.5 times higher risk of atypical profile than Swedish Red and Whites. Risk of atypical profiles was 0.5 and 0.7 times lower for older cows compared with first-parity cows; 2.3 times higher for cows in tie-stalls compared with those in loose housing; 2.6 times higher for cows calving during winter compared with summer; 0.5 times lower for cows in earlier (1994-1999) calving-year groups compared with the most recent (2000-2002); 2.5 times higher for cows with planned extended calving interval compared with conventional calving interval; and 2.2 times higher for an atypical profile in previous lactation compared with a normal profile. Cows with atypical profiles had a 15-d increase in interval from calving to first artificial insemination and an 18-d increase in interval from calving to conception. Progesterone samples taken within the first 60 d postpartum were used to calculate the percentage of samples above the threshold value of luteal activity. This measure had a significantly different mean in profiles and can be used to separate delayed onset of cyclical ovarian activity profiles and prolonged luteal phase profiles from normal. Thereby, it may be a more effective tool than measurements based only on the onset of ovarian cyclical activity in genetic evaluation of early postpartum fertility in dairy cows.