Blood metabolites and hormone-based programmed breeding treatments in anovular lactating dairy cows

The objectives were to (1) compare blood metabolites and reproductive outcomes in lactating dairy cows not inseminated before (early) and after (late) 100 d in milk (DIM) because of prolonged anovulation or anestrus; and (2) evaluate reproductive responses of cows ≤100 DIM to GnRH + PGF_2α treatments after a fixed-time artificial insemination (AI; Ovsynch) or after induced estrus (Select Synch). In blood samples collected before initiating hormone-based breeding programs, anovular cows ≤100 DIM had the greatest serum total protein and globulin concentrations and the lowest tri-iodothyronine concentrations. Anovular and ovular cows >100 DIM had the greatest serum urea concentrations. Ovaries in cows (n = 40) >100 DIM were examined by transrectal ultrasonography, and those without a detectable corpus luteum (CL; anovular) were given GnRH and then PGF_2α 7 d later (Select Synch), whereas cows with a CL (ovular) were given 2 PGF_2α injections 12 d apart. Cows were inseminated at observed estrus after the second or only PGF_2α injection. More ovular (79%; 15/19) than anovular cows (24%; 5/21) were detected in estrus. No differences were detected between ovular and anovular cows in DIM at first AI, conception rate to first AI, cumulative pregnancy rates, number of services per conception, or days open. Cows (n = 93) ≤100 DIM were assigned randomly to 3 treatments: (1) control (n = 20) AI at estrus; (2) GnRH and then PGF_2α on 7 d (Select Synch; n = 42) and monitored for signs of estrus for 5 d and AI accordingly; or (3) 2 GnRH injections 9 d apart with PGF_2α given 48 h before second GnRH injection and AI at 16 h after the second GnRH injection (Ovsynch; n = 31). Among cows ≤100 DIM, controls had more days to first service (149 ± 16 d) than Select Synch cows (117 ± 7 d). Ovsynch cows had the fewest days to first service (84 ± 10 d) compared with control (149 ± 16 d) and Select Synch (117 ± 7 d) cows. Conception rates in control (25%) and Select Synch (26.2%) cows did not differ from those in Ovsynch cows (29%). Ovsynch cows had greater cumulative pregnancy rates and fewer days open than control (161 ± 20 vs. 258 ± 29 d), but did not differ from Select Synch (233 ± 19 d). Timed AI produced comparable fertility and superior cumulative pregnancy rates, fewer days to first service, and fewer days open than AI at observed estrus in cows inseminated ≤100 DIM.     

Prevalence and risk factors related to anovular phenotypes in dairy cows

The objective of the current study was to evaluate the relationship of body condition score (BCS) at 35 d in milk (DIM), milk production, diseases, and duration of the dry period with prevalence of anovulation at 49 DIM and then, specifically, with the prevalence of each anovular phenotype. We hypothesized that anovular follicular phenotypes, classified based on maximal size of the anovular follicle, have different etiologies. A total of 942 lactating Holstein cows (357 primiparous and 585 multiparous) from 1 herd had ovaries evaluated by ultrasonography at 35 ± 3 and 49 ± 3 DIM to detect the absence of a corpus luteum (CL), and to measure the diameter of the largest follicle. Cows were classified as cyclic at 49 DIM if a CL was observed in at least 1 of the 2 examinations, or anovular if no CL was observed at either examination. Cows considered anovular were divided into 3 groups based on the largest diameter of the largest follicle as follows: ranging from 8 to 13 mm, 14 to 17 mm, or ≥18 mm. Cows were evaluated for the following diseases: retained placenta, metritis, hyperketonemia, mastitis, lameness, respiratory problem, and digestive problem. At 35 DIM, BCS was determined, and milk yield for individual cows was recorded. A total of 28.5% (268/942) of cows were classified as anovular. Anovular cows had longer dry periods (90 vs. 71 d) and smaller BCS than cyclic cows (2.83 vs. 2.99). Cows with a single disease or multiple diseases had 2 and 3-fold increase in odds of being anovular, respectively. Anovular cows had follicles that ranged from 4 to 50 mm. The prevalence of anovular phenotype, among anovular cows, that had the diameter of the largest follicle ranging from 8 to 13 mm, 14 to 17 mm, and ≥18 mm was 29.9 (79/264), 37.5 (99/264), and 32.6% (86/264), respectively. Anovular cows with follicles of 8 to 13 mm had longer dry periods than those with follicles ≥18 mm (104 vs. 74 d), whereas anovular cows with medium size follicles had intermediate days dry (99 d). Cows with small and medium anovular follicles had smaller BCS and greater prevalence of multiple diseases than cyclic cows. For almost all risk factors, the cows with large anovular follicles (≥18 mm) were similar to cyclic cows and different from cows with smaller anovular follicles (8–13 mm). Thus, longer dry periods, less BCS at 35 DIM, and diseases were risk factors for anovulation. Moreover, the risk factors for the 3 distinct anovular follicle phenotypes differed.     

Stage of cycle, incidence, and timing of ovulation, and pregnancy rates in dairy cattle after three timed breeding protocols

In experiment 1, 705 cows were assigned to three treatments: 1) the Ovsynch protocol (a GnRH injection given 7 d before and another 48 h after one PGF2alpha injection); 2) PGF2alpha + Ovsynch (one PGF2alpha injection given 12 d (d -22) before initiating Ovsynch (d -10); and 3) 2xPG12 (two PGF2alpha injections 12 d apart; d -15 and -3, followed 48 h later by a GnRH injection. All cows were inseminated (d 0) 16 to 20 h after the GnRH injection on d -1. Cyclic status was estimated by serum progesterone. More cows were in early diestrus at d -10 for PGF2alpha + Ovsynch (36%) and 2 x PG12 (29%) versus Ovsynch (19%). Multiparous cows receiving PGF2alpha + Ovsynch had greater pregnancy rates via ultrasonography at d 28 after AI (42%) than contemporaries after Ovsynch (28%) or 2xPG12 (27%) but did not differ significantly at palpation 10 to 30 d later (28, 19, and 17%, respectively). Pregnancy of first-parity cows was similar across treatments at 28 d (41%) or at palpation (33%). Pregnancy rates for 128 anestrous cows were lower, regardless of treatment. Overall embryo survival from d 28 until palpation was 72% but was only 44% in 2xPG12 cows that were anestrus through d -10. Experiment 2 included the three treatments above plus controls (one GnRH injection 7 d before PGF2alpha and AI after estrus). Preovulatory follicles were 6 to 11% larger near estrus in controls than on d -1 in cows receiving GnRH. More controls ovulated by 32 h after onset of estrus than were treated cows by 32 h after GnRH, but percentages (79 to 94%) were similar by 40 h. In multiparous cows, PGF2alpha before Ovsynch increased pregnancy rates, whereas the 2xPG12 protocol produced similar pregnancy rates as Ovsynch across parities. Ovulation was effectively induced by 40 h after GnRH.     

Follicular wave of the ovulatory follicle and not cyclic status influences fertility of dairy cows

Two experiments evaluated the influence of follicular wave at artificial insemination (AI) on fertility of dairy cows. In experiment 1, data from 5,607 lactating cows enrolled in estrous and ovulation synchronization programs for AI were evaluated. Cows’ blood was analyzed for progesterone 7 to 14 d apart, with the second sample collected on the day of the first GnRH (GnRH1) of the synchronization protocol. Cows were classified as cyclic if progesterone was ≥1 ng/mL in at least 1 of the 2 samples and as anovular if both samples were <1 ng/mL. Cyclic cows were categorized as low (CLOW; < 1 ng/mL) or high (CHIGH; ≥ 1 ng/mL) progesterone on the day of GnRH1, which would result in ovulation of the dominant follicle of the first (FW) and second (SW) follicular waves, respectively, at AI. Pregnancy per AI (P/AI) was determined 30 and 53 d after AI. In experiment 2, 220 cyclic Holstein cows received 2 injections of PGF_2α administered 14 d apart. The Ovsynch protocol (d 0 GnRH, d 7 PGF_2α, d 9 GnRH, d 9.5 timed AI) was initiated either 3 or 10 d after the second PGF_2α of the presynchronization to result in insemination to the FW or SW dominant follicles. Blood was analyzed for progesterone and ovaries were scanned to determine ovulatory responses and follicle diameter. Pregnancy was determined on d 32 and 67 after timed AI. In experiment 1, P/AI on d 30 was greater for CHIGH cows than for CLOW and anovular cows (43.0, 31.3, and 29.7%, respectively), but because of pregnancy loss, P/AI on d 53 was lowest for anovular cows. Proportions of cows with short reinsemination intervals differed among groups and were 7.1, 15.7, and 11.9% for CHIGH, CLOW, and anovular cows, respectively. Pregnancy loss was greater for anovular cows than for CLOW cows (15.0 vs. 10.0%) and was intermediate for CHIGH cows (13.5%). In experiment 2, 9.8 and 97.2% of the FW and SW cows, respectively, had progesterone ≥1 ng/mL at GnRH1. Concentrations of progesterone at the GnRH1 and PGF_2α injections of the Ovsynch protocol were greater for SW cows than FW cows. Pregnancy per AI was greater for SW cows than for FW cows (41.7 vs. 30.4%) despite less ovulation to GnRH1 in SW cows than in FW cows (78.7 vs. 88.4%). Collectively, these data indicate that follicular wave of the ovulatory follicle and not cyclic status caused the greatest reduction in P/AI in dairy cows. Whether the culprit is the follicle itself or the hormonal milieu characteristic of the first follicular wave and the early stage of the estrous cycle remains to be elucidated. Synchronization programs that induced ovulation of the FW follicle at AI reduced P/AI in lactating dairy cows, and ovulation of the FW follicle, or development of the ovulatory follicle under low progesterone concentrations, or both, might be mechanisms for reduced fertility in anovular cows.     

Relationship between level of milk production and multiple ovulations in lactating dairy cows

Our objective was to evaluate factors associated with spontaneous multiple ovulations in lactating dairy cows. Ovaries of cows [n = 267; >50 days in milk (DIM)] were evaluated weekly using ultrasound to determine spontaneous (i.e., no hormonal treatment) ovulation rate starting at 50 DIM and continuing until pregnancy diagnosis. Cows were fitted with a transmitter to record standing activity during estrus, and serum progesterone concentration was assessed weekly starting at wk 1 postpartum for all cows. Overall, 76 (28.5%) cows were anovular and 191 (71.5%) were ovular by 71 DIM. Incidence of anovulation was not associated with level of milk production but was associated with lower body condition. For anovular cows (n = 41) that spontaneously recovered, the multiple ovulation rate at first ovulation was 46.3%. For second and subsequent ovulations (n = 463), the level of milk production for 14 d preceding estrus was associated with increased ovulation rate. To illustrate, incidence of multiple ovulations was 1.6% (2/128), 16.9% (32/189), and 47.9% (70/146) for ovulations when cows were producing <35, 35 to <45, and ≥45 kg/d, respectively. Among cows for which estrous behavior was recorded, those with multiple ovulations (n = 48) had shorter duration of estrus (4.3 ± 0.7 vs. 9.9 ± 0.5 h) and higher production (47.2 ± 0.9 vs. 38.1 ± 0.5 kg/d) than cows with single ovulations (n = 237). Circulating concentrations of estradiol were lower (5.5 ± 0.3; n = 15 vs. 7.8 ± 0.4 pg/mL; n = 71) during periods of estrus with multiple ovulations despite a greater preovulatory follicular volume (4136 ± 123 vs. 3085 ± 110 mm³). Similarly, serum progesterone concentration 7 d after estrus was lower for cows with multiple than single ovulations (2.5 ± 0.3 vs. 3.2 ± 0.1 ng/mL) despite a greater luteal volume (8291 ± 516 vs. 6405 ± 158 mm³). In summary, the first spontaneous ovulation in anovular cows and a higher level of milk production for 14 d preceding estrus were associated with increased multiple ovulation rate. Additionally, cows with multiple ovulations had lower estradiol at estrus, a shorter duration of estrus, and lower progesterone at 7 d after estrus than cows with single ovulations.     

Supplementation with estradiol-17beta before the last gonadotropin-releasing hormone injection of the Ovsynch protocol in lactating dairy cows

The aim of this study was to determine whether an increase in circulating estrogen concentrations would increase percentage pregnant per artificial insemination (PP/AI) in a timed AI protocol in high-producing lactating dairy cows. We analyzed only cows having a synchronized ovulation to the last GnRH of the Ovsynch protocol (867/1,084). The control group (n = 420) received Ovsynch (GnRH - 7 d - PGF_2α - 56 h - GnRH - 16 h - timed AI). The treatment group (n = 447) had the same timed AI protocol with the addition of 1 mg of estradiol-17β (E2) at 8 h before the second GnRH injection. Ovarian ultrasound and blood samples were taken just before E2 treatment of both groups. In a subset of cows (n = 563), pressure-activated estrus detection devices were used to assess expression of estrus at 48 to 72 h after PGF_2α treatment. Ovulation was confirmed by ultrasound 7 d after timed AI. Treatment with E2 increased expression of estrus but overall PP/AI did not differ between E2 and control cows. There was an interaction between treatment and expression of estrus such that PP/AI was greater in E2-treated cows that showed estrus than in E2-treated or control cows that did not show estrus and tended to be greater than control cows that showed estrus. There was evidence for a treatment by ovulatory follicle size interaction on PP/AI. Supplementation with E2 improved PP/AI in cows ovulating medium (15 to 19 mm) but not smaller or larger follicles. The E2 treatment also tended to improve PP/AI in primiparous cows with low (≤2.5) body condition score, and in cows at first postpartum service compared with Ovsynch alone. In conclusion, any improvements in PP/AI because of E2 treatment during a timed AI protocol appear to depend on expression of estrus, parity, body condition score, and size of ovulatory follicle.     

Prostaglandin F2α and gonadotropin-releasing hormone administration improve progesterone status, luteal number, and proportion of ovular and anovular dairy cows with corpora lutea before a timed artificial insemination program

The objective of this research was to increase the proportion of cows with at least 1 functional corpus luteum (CL) and elevated progesterone at the onset of the timed artificial insemination (TAI) protocol. Postpartum Holstein cows in one herd were stratified by lactation number at calving (September 2009 through August 2010) and assigned randomly to 2 treatments: 1) Presynch-10 (n=105): two 25-mg injections of PGF(2α) (PG) 14 d apart (Presynch); and 2) PG-3-G (n=105): one 25-mg injection of PG 3 d before 100-μg GnRH (Pre-GnRH) injection, with the PG injection administered at the same time as the second PG injection in the Presynch-10 treatment. Cows were enrolled in a TAI protocol [Ovsynch; injection of GnRH 7 d before (GnRH-1) and 56 h after (GnRH-2) PG injection with AI 16 to 18 h after GnRH-2] 10 d after the second or only PG injection. Blood samples for progesterone or estradiol analyses were collected on median days in milk (DIM): 36, 39, 50, 53 (Pre-GnRH), 60 (GnRH-1), 67 (PG), 69 (GnRH-2), and 70 (TAI). Ovarian structures were measured by ultrasonography on median DIM 53, 60, 67, 69, and 6 d post-TAI to determine follicle diameters, ovulation response to GnRH, or both. Although progesterone concentration did not differ between treatments before Pre-GnRH injection, the proportion of cows with at least 1 CL tended to be greater for PG-3-G than Presynch-10 cows, and more PG-3-G cows ovulated after Pre-GnRH injection than ovulated spontaneously in Presynch-10. Further, the diameter of follicles that ovulated tended to be smaller in PG-3-G than in Presynch-10 cows after Pre-GnRH injection. At GnRH-1, the proportion of cows with progesterone ≥1 ng/mL, the number of CL per cow, and the proportion of cows with at least 1 CL were greater for PG-3-G than Presynch-10. Neither follicle diameter nor percentage of cows ovulating after GnRH-1 differed between treatments. At PG injection during the week of TAI, progesterone concentration and the proportion of cows with progesterone ≥1 ng/mL tended to be greater for PG-3-G than Presynch-10, and PG-3-G had more CL per cow than Presynch-10. No ovarian characteristics differed between treatments after GnRH-2, including progesterone concentration, number of CL per cow, and total luteal volume 7 d after GnRH-2. Many of the previous ovarian traits were improved in both ovular and anovular cows after PG-3-G compared with Presynch-10. Pregnancies per AI at d 32 and 60 were only numerically greater for PG-3-G than for Presynch-10 cows, largely because of differences detected during months without heat stress. We concluded that the PG-3-G treatment increased ovulation rate and luteal function 7 d before the onset of Ovsynch, resulting in improved follicular synchrony and predisposing potentially greater pregnancies per AI in lactating dairy cows.     

Effect of bovine somatotropin (500 mg) administered at ten-day intervals on ovulatory responses, expression of estrus, and fertility in dairy cows

The objectives of this study were to evaluate the effect of administering 500 mg of recombinant bovine somatotropin (bST) every 10 d on ovulatory responses, estrous behavior, and fertility of lactating Holstein cows. Lactating dairy cows were assigned to 1 of 2 treatments: a control with no administration of bST (73 primiparous and 120 multiparous cows) or 6 consecutive administrations of 500 mg of bST (83 primiparous and 123 multiparous cows) given subcutaneously at 10-d intervals starting 61 ± 3 d postpartum (study d 0), concurrent with the initiation of the timed artificial insemination (AI). Blood samples were collected thrice weekly from 61 ± 3 to 124 ± 3 d in milk (DIM), and plasma samples were analyzed for concentrations of estradiol, glucose, insulin, insulin-like growth factor 1, and progesterone. The estrous cycle of cows was presynchronized with 2 injections of PGF_2α at 37 ± 3 and 51 ± 3 DIM, and the Ovsynch timed AI protocol was initiated at 61 ± 3 DIM. Ovaries were scanned to determine ovulatory responses during the Ovsynch protocol. Pregnancy was diagnosed at 33 and 66 d after AI. Body condition was scored on study d 0, 10, 42, and 76. Sixty-four cows were fitted with a pressure mounting sensor with radiotelemetric transmitters to monitor estrous behavior. Treatment of lactating dairy cows with 500 mg of bST at 10-d intervals increased yields of milk and milk components in the first 2 mo after treatment. Body condition of bST-treated cows remained unaltered, whereas control cows gained BCS. Treatment with bST increased concentrations of insulin-like growth factor 1 chronically, but concentrations of insulin and glucose increased only transiently in the first 7 d after the first injection of bST. Concentrations of progesterone during and after the Ovsynch protocol remained unaltered after treatment with bST; likewise, ovulatory responses during the Ovsynch protocol were mostly unaltered by treatment. Concentration of estradiol tended to be greater for bST cows than for control cows immediately before induction of ovulation in the Ovsynch protocol. Similarly, the mean and the peak concentrations of estradiol were greater for bST cows than for control cows when monitored during spontaneous estrus. Nevertheless, duration of estrus and the median number of standing events were less for bST cows than for control cows. Pregnancies per AI after the first and second postpartum inseminations were not affected by bST treatment. Treatment of lactating dairy cows with 500 mg of bST every 10 d improved lactation performance, but it did not affect pregnancies per AI and it reduced expression of estrus.     

Short communication: A positive relationship between the first ovulation postpartum and the increasing ratio of milk yield in the first part of lactation in dairy cows

The aim of the present study was to examine the relationship between characteristics of the lactation curve, on the basis of daily milk yield, and ovulation within 3 wk postpartum as an indicator of early return to luteal activity in dairy cows. Lactation records from 46 lactating Holstein cows between calving and 305 d postpartum were studied. Milk samples were collected twice weekly between d 7 and 100 for later determination of progesterone concentrations. Occurrence of an early first ovulation was determined by an increase in milk progesterone by 3 wk after calving. Milk yield was recorded daily until 305 d postpartum, and average yield was calculated weekly. The lactation curve was characterized by 8 indices on the basis of the weekly average of milk yield as follows: a) first-week milk yield; b) peak milk yield; c) actual 305-d milk yield; d) peak week; e) difference in milk yield between the first week and peak week; f) difference in milk yield between the peak week and last week (43rd week postpartum); g) ratio of increase in milk yield between wk 1 and the week of peak yield; and h) ratio of decline in milk yield between the week of peak yield and the last week. Indices g and h were calculated as linear. The number of cows having ovulated by 3 wk postpartum was 22 (47.8%). The resumption of ovarian cycles with normal luteal phases occurred earlier in ovular cows than in anovular cows (32.0 d vs. 57.1 d). Although total milk yield did not differ between ovular and anovular cows, the ratio of increase in milk yield from the first week to the peak week (index g) in ovular cows was smaller compared with that of anovular cows (1.71 vs. 2.54). In addition, the ratio of increase in milk yield from the first week to the third week postpartum was greater in anovular cows by 3 wk postpartum (ovular = 1.43 ± 0.23 vs. anovular = 2.32 ± 0.29). In conclusion, the present study demonstrates that a greater increasing ratio of milk yield during early lactation may delay resumption of ovarian cycles after parturition. Therefore, this study is the first to demonstrate statistically that a smaller increasing ratio of milk yield (index g) during early lactation may have a beneficial effect on the first ovulation by 3 wk postpartum.     

Resynchronizing estrus and ovulation after not-pregnant diagnosis and various ovarian states including cysts

We compared outcomes of 2 protocols used to resynchronize estrus and ovulation in dairy females after a not-pregnant diagnosis. Nulliparous heifers and lactating cows in which artificial insemination (AI) occurred 41 +/- 1 d earlier were presented every 2 to 3 wk for pregnancy diagnosis by using ultrasonography. Ovaries were scanned, follicles were mapped and sized, presence of corpus luteum was noted, and GnRH was injected (d 0). Females were assigned randomly to receive PGF(2alpha) 7 d later (d 7) and then either received estradiol cypionate (ECP) 24 h after PGF(2alpha) (d 8; Heatsynch; n = 230) or a second GnRH injection 48 h after PGF(2alpha) (d 9; Ovsynch; n = 224). Those detected in estrus since their not-pregnant diagnosis were inseminated, whereas the remainder received a timed AI (TAI) between 65 and 74 h after PGF(2alpha). Ovarian scans and blood collected before injections for progesterone analysis were used to classify 4 ovarian status groups: anestrus, follicular cysts, luteal cysts, and cycling, plus an unknown group of females in which no blood sampling or ovarian scans were made. Few females (5.1%) were inseminated between not-pregnant diagnosis and d 8. On d 10, more ECP- than GnRH-treated females were inseminated after detected estrus (24 vs. 6%). Overall, more Ovsynch than Heatsynch females received a TAI (82 vs. 62%). Conception rates tended to be greater for females inseminated after estrus (37%) than after TAI (29%), particularly for those treated with Heatsynch (41 vs. 27%) than with Ovsynch (33 vs. 31%). Those inseminated after estrus conceived 31 +/- 8 d sooner than those receiving the TAI. Conception rates for females having elevated progesterone 7 d after the not-pregnant diagnosis were greater than those having low progesterone in Heatsynch (42%; n = 133 vs. 25%; n = 55) and Ovsynch protocols (33%; n = 142 vs. 15%; n = 45), respectively. Conception rates were greater in nulliparous heifers than in lactating cows (43 vs. 28%) regardless of protocol used. Although overall pregnancy outcomes after a not-pregnant diagnosis were similar in response to either the Ovsynch and Heatsynch protocols, inseminations performed after detected estrus before the scheduled TAI reduced days to eventual conception and tended to increase conception rates, particularly after Heatsynch.     

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.     

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.     

Inseminations at estrus induced by presynchronization before application of synchronized estrus and ovulation

A controlled field study examined conception rates after 2 timed artificial insemination (TAI) breeding protocols conducted on 2 commercial dairy farms. Estrous cycles in postpartum lactating cows were presynchronized with 2 injections of PGF(2alpha) given 14 d apart (Pre-synch) and then, after 12 d, the standard Ovsynch protocol (injection of GnRH 7 d before and 48 h after an injection of PGF(2alpha), with one TAI at 12 to 16 h after the second GnRH injection) or Heatsynch protocol [injection of GnRH 7 d before an injection of PGF(2alpha), followed 24 h later by 1 mg of estradiol cypionate (ECP) and one TAI 48 h after ECP] was applied. Experimental design allowed artificial insemination to occur anytime after the second Presynch injection and during the designed breeding week when estrus was detected. Of the 1846 first services performed, only 1503 (rate of compliance = 81.4%) were performed according to protocol. Numbers of cows inseminated, logistic-regression adjusted conception rates, and days in milk (DIM) were for inseminations made: 1) during 14 d after first Presynch injection (n = 145; 22.6%; 54 +/- 0.4 DIM); 2) during 12 d after second Presynch injection (n = 727; 33%; 59 +/- 0.2 DIM); 3) during 7 d after the first GnRH injection of Ovsynch or Heatsynch (n = 96; 32.1%; 74 +/- 0.5 DIM); 4) after estrus as part of Heatsynch (n = 212; 44.6%; 76 +/- 0.3 DIM); 4) after TAI as part of Heatsynch (n = 154; 21.1%; 76 +/- 0.4 DIM); 5) after estrus as part of Ovsynch (n = 43; 48.7%; 77 +/- 0.7 DIM); and 6) after TAI as part of Ovsynch (n = 271; 24.4%; 77 +/- 0.3 DIM). Conception rates when AI occurred after one Presynch injection were less than when AI occurred after 2 Presynch injections. Conception rates for those inseminated after either Presynch injection did not differ from those inseminated after combined Heatsynch + Ovsynch. Cows in the Ovsynch and Heatsynch protocols inseminated after estrus during the breeding week had greater conception rates than those receiving the TAI, but overall conception rates did not differ between protocols. Among cows inseminated after detected estrus, conception was greater for cows in the Heatsynch + Ovsynch protocol (77 +/- 0.4 DIM) than for those inseminated after either Presynch injection (54 +/- 0.4 or 59 +/- 0.2 DIM). We concluded that conception rates after Heatsynch and Ovsynch were similar under these experimental conditions, and that delaying first AI improved fertility for cows inseminated after detected estrus.     

Evaluation of two hormonal protocols for synchronization of ovulation and timed artificial insemination in dairy cows managed in grazing-based dairies

To evaluate the efficacy of two hormonal protocols for synchronization of ovulation and timed artificial insemination (TAI) in dairy cows managed in grazing-based dairies, lactating dairy cows (n = 142) from two grazing-based dairies were randomly assigned to one of three treatment groups. Cows in the first group (Ovsynch) received 50 microg of GnRH (d -10); 25 mg of PGF2alpha (d -3), and 50 microg of GnRH (d -1) followed by timed AI on d 0. Cows in the second group (PGF + Ovsynch) received a modified Ovsynch and timed AI similar to Ovsynch but with the addition of 25 mg of PGF2alpha 12 d (d -22) before initiation of Ovsynch. Cows in the third group (control) received standard reproductive management in place on each farm. Luteolysis occurred in 90.5% of cows exhibiting luteal function on d -22 in the PGF + Ovsynch treatment group, whereas none of the cows in the Ovsynch group underwent luteolysis on d -22. Synchronization rate (i.e., ovulatory response at 48 h after the second GnRH injection), conception rates at TAI and pregnancy rates after 35 d of breeding were similar for cows in the Ovsynch and PGF + Ovsynch groups. The proportion of anovular cows at the first GnRH injection of the synchronization protocols (d -10) was similar for cows receiving Ovsynch (28.0%) and PGF + Ovsynch (30.7%), and conception rate at TAI was similar for cycling (45.8%) and anovular (30.0%) cows. The cumulative pregnancy rate was greater for cows receiving TAI compared with control cows after 7 d of breeding (41.2 vs. 20.0%) but did not differ at 35 d of breeding (54.9 vs. 60.0%). Administration of PGF2alpha 12 d before initiation of Ovsynch did not improve synchronization, conception, or pregnancy rate compared with the standard Ovsynch protocol. Synchronization of ovulation to initiate timed AI at the onset of the breeding season resulted in earlier establishment of pregnancy compared with standard reproductive management.     

Comparison of responses to Ovsynch between Holstein-Friesian and Swedish Red cows

The Ovsynch protocol was designed to synchronize ovulation, thereby allowing timed artificial insemination (TAI) of all cows without detection of estrus. However, the effectiveness of Ovsynch in different breeds of dairy cows has not been previously compared. The aim of this study was to compare the response to Ovsynch in cycling lactating Holstein-Friesian (HF) and Swedish Red (SR) dairy cows. A total of 495 cyclic cows (n = 347 HF, n = 148 SR) were housed together and treated with Ovsynch (GnRH - 7 d - PGF_2α - 56 h - GnRH - 16 to 18 h - TAI). Ovulatory responses, synchronization rate, maximal follicle size at the time of AI, and percentage of pregnant cows per AI (P/AI at 31 and 62 d after AI) were compared between breeds. Ultrasonography was performed during Ovsynch at first GnRH, PGF_2α, at time of AI, and 7 d after AI. Ovulatory response and synchronization rate were similar in HF versus SR cows (60.2 vs. 62.2%; 88.4 vs. 88.5%, respectively). Cows that ovulated to the first GnRH of Ovsynch had smaller follicle size at AI (15.9 ± 0.1 vs. 16.4 ± 0.2 mm). Maximal follicle size at AI was greater for HF (16.4 ± 2.2 mm) than SR (15.5 ± 2.3 mm) cows. The P/AI was greater for SR than HF cows at the 62-d pregnancy diagnosis (56.1 vs. 46.1%). In addition, pregnancy loss between 31 and 62 d of pregnancy was greater in HF (10.1%) than SR (3.5%) cows. Fertility was less in HF cows during the hot season (57.7 in cold vs. 38.1% in the hot season), whereas such a decrease was not observed in SR (60.0 in cold vs. 53.5% in the hot season) cows. Thus, although the GnRH treatments of Ovsynch were equally effective in SR and HF cows, pregnancy outcomes (P/AI at d 62 and pregnancy survival) were greater in SR than HF cows, and P/AI in SR cows was not compromised during the hot season as was found for HF cows.     

Presynchronization with prostaglandin F2α and gonadotropin-releasing hormone simultaneously improved first-service pregnancy per artificial insemination in lactating Holstein cows compared with Presynch-14 when combined with detection of estrus

This study aimed to determine the effect of 2 simple breeding strategies combining artificial insemination (AI) after detection of estrus (AIED) and timed AI (TAI) on first-service fertility in lactating Holstein cows. Weekly, lactating Holstein cows (n = l,049) between 40 and 46 d in milk (DIM) were randomly assigned to initiate 1 of 2 breeding strategies for first service: Presynch-14 and PG+G. Presynch-14 is a presynchronization strategy with 2 PGF_2α treatments 14 d apart with the last PGF_2α 14 d before the initiation of the Ovsynch protocol. Cows treated with PG+G receive a simpler presynchronization program that uses PGF_2α and GnRH simultaneously 7 d before Ovsynch. In both treatments, cows detected in standing estrus by tail chalk at any time ≥55 DIM were inseminated, and treatment was discontinued (n = 525). Cows completing treatment received TAI from 78 to 84 DIM (n = 526). In a subgroup of cows that received TAI, blood was collected (n = 163) to assess circulating concentrations of progesterone, and ultrasonographic evaluations of ovaries were performed on the day of first GnRH of Ovsynch (n = 162) and PGF_2α of Ovsynch (n = 122). The proportion of cows that received TAI was greater for PG+G compared with Presynch-14 (63.5 vs. 31.9%), which increased DIM at first service for cows treated with PG+G compared with Presynch-14 (75.5 ± 0.4 vs. 68.7 ± 0.4). For cows receiving TAI, the ovulatory response to first GnRH of Ovsynch (73.8 vs. 48.8%) and the proportion of cows with functional corpora lutea (92.6 vs. 73.1%) were greater for PG+G than Presynch-14. Cows treated with PG+G had greater overall pregnancy per AI (P/AI) 42 ± 7 d after AI (40.2 vs. 33.6%) and calving per AI (32.1 vs. 25.2%) than Presynch-14. For cows receiving AIED, treatment did not affect P/AI 42 ± 7 d after AI. However, for cows receiving TAI, PG+G increased P/AI compared with Presynch-14 (44.6 vs. 35.2%). Overall, cows receiving TAI had greater P/AI 42 ± 7 d after AI (42.5 vs. 31.5%) and calving per AI (34.1 vs. 23.7%) and decreased pregnancy loss (16.8 vs. 25.2%) than cows receiving AIED. In summary, PG+G increased the proportion of cows receiving TAI and the DIM at first service, P/AI, and calving per AI compared with Presynch-14 when both TAI programs were combined with AIED.     

Assessment of an accelerometer system for detection of estrus and treatment with gonadotropin-releasing hormone at the time of insemination in lactating dairy cows

Two experiments were conducted to evaluate an accelerometer system (Heatime; SCR Engineers Ltd., Netanya, Israel) to manage reproduction in lactating dairy cows. In experiment 1, lactating Holstein cows (n = 112) were fitted with an accelerometer system and were treated with GnRH followed 7 d later by PGF_2α to synchronize estrus. A total of 89 cows that had a follicle >10 mm in diameter and a functional corpus luteum at the PGF_2α injection that regressed by 48 h after induction of luteolysis were included in the analysis. Overall, 71% of cows were detected in estrus by the accelerometer system and 95% of cows showing estrus ovulated within 7 d after induction of luteolysis. Of the cows not detected in estrus by the accelerometer system, 35% ovulated within 7 d after induction of luteolysis. Duration of estrus activity (mean ± SD) was 16.1 ± 4.7 h and was neither affected by parity nor milk production. Intervals (means ± SD) from induction of luteolysis, onset of activity, peak raw activity, and peak weighted activity to ovulation was 82.2 ± 9.5, 28.7 ± 8.1, 20.4 ± 7.8, and 16.4 ± 7.4 h, respectively, and the interval from AI to ovulation was 7.9 ± 8.7 h, but ranged from −12 to 26 h. In experiment 2, cows were assigned randomly to receive an intramuscular injection of GnRH at artificial insemination (AI) after detection of estrus by the accelerometer system or receive no treatment (control). Nine hundred seventy-nine AI services from 461 cows were analyzed. Treatment with GnRH at AI did not affect fertility at 35 or 65 d after AI, and no interaction was detected between treatment and season or treatment and AI number. Overall, two-thirds of the cows that were considered properly synchronized were inseminated based on the accelerometer system and ovulated after AI. The remaining cows either were not inseminated because they were not detected in estrus or would not have had a chance to conceive to AI because they failed to ovulate after estrus. Furthermore, mean time of AI in relation to ovulation determined by the accelerometer system was acceptable for most of the cows that displayed estrus; however, variability in the duration of estrus and timing of AI in relation to ovulation could lead to poor fertility in some cows. For lactating dairy cows detected in estrus by the accelerometer system, treatment with GnRH at the time of AI without reference to the onset of estrus did not increase fertility.     

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.     

An alternative AI breeding protocol for dairy cows exposed to elevated ambient temperatures before or after calving or both

Our objective was to determine if a timed artificial insemination (AI) protocol (Ovsynch) might produce greater pregnancy rates than AI after a synchronized, detected estrus during summer. Lactating Holstein cows (n = 425) were grouped into breeding clusters and then assigned randomly to each of two protocols for AI between 50 and 70 days in milk. All cows were treated with GnRH followed 7 d later by PGF2alpha. Ovsynch cows then were treated with a second injection of GnRH 48 h after PGF2alpha and inseminated 16 to 19 h later. Controls received no further treatment after PGF2alpha and were inseminated after detected estrus. Pregnancy was diagnosed once by transrectal ultrasonography (27 to 30 d after AI) and again by palpation (40 to 50 d). Based on concentrations of progesterone in blood collected before each hormonal injection, only 85.4% of 425 cows were considered to be cycling. Although conception rates were not different between protocols at d 27 to 30, AI submission rates and pregnancy rates were greater after Ovsynch (timed AI) than after detected estrus. A temperature-humidity index > or = 72 was associated with fewer controls detected in estrus with lower conception than for controls detected in estrus when index values were < 72, whereas the reverse was true for cows after the Ovsynch protocol. We concluded that a timed AI protocol increased pregnancy rates at d 27 to 30 because its success was independent of either expression or detection of estrus. However, because of poorer embryonic survival in Ovsynch cows during heat stress only (39.5 vs. 69.2% survival for Ovsynch and control, respectively), pregnancy rates were not different by d 40 to 50 after timed AI.     

Initiation of the breeding season in a grazing-based dairy by synchronization of ovulation

Lactating dairy cows (n = 228) in a semiseasonal, grazing-based dairy were subjected to artificial insemination (AI) to start the 23-d breeding season (d 0 to 22) followed by natural service (d 23 to 120). Cows were randomly assigned to: 1) Ovsynch (GnRH, d -10; PGF2,, d -3; GnRH, d -1; timed AI, d 0) followed by AI at estrus (tail paint removal) on d 1 to 22 (Ovsynch; n = 114); or 2) AI at estrus (tail paint removal) throughout 23 d of AI breeding (tail paint; n = 114). Days to first AI service were greater and the 23-d AI service rate was less for tail paint vs. Ovsynch cows (12.0 +/- 0.6 d vs. 0 d; and 84.2 vs. 100%, respectively). However, conception to first AI was greater for tail paint vs. Ovsynch cows (47.3 vs. 27.3%, respectively). Cows in the tail paint group received only one AI, during 23 d of AI, but 46.4% of Ovsynch cows received a second AI, with similar conception (43.1%) to that of tail paint cows at first AI (47.3%). Based on serum progesterone, incomplete luteal regression after PGF2alpha, and poor ovulatory responses to GnRH contributed to lower conception to timed AI in the Ovsynch group. Cumulative pregnancy rates for tail paint and Ovsynch cows did not differ after 23 d of AI breeding (47.3 vs. 46.3%, respectively) nor after 120 d of AI/ natural service breeding (80.5 vs. 83.3%, respectively). Lactating cows in this grazing-based dairy synchronized poorly to Ovsynch resulting in reduced conception to timed AI compared with AI after tail paint removal.