M. F. Sá Filho

Risk factors for resumption of postpartum estrous cycles and embryonic survival in lactating dairy cows

The objectives of this study were to evaluate factors associated with resumption of postpartum estrous cycles and embryonic survival in lactating dairy cows. Holstein cows, 6396 from four dairy farms were evaluated to determine the relationships among parity, body condition score (BCS) at calving and at AI, season of year when cows calved, and milk yield on resumption of postpartum estrous cycles by 65 days postpartum, and all the previous variables, estrual or anestrus and AI protocol on conception rates and embryonic survival at the first postpartum insemination. Cows had their estrous cycle pre-synchronized with two PGF(2alpha) injections given 14 days apart and were inseminated between 69 and 82 days postpartum following either an estrous or ovulation synchronization protocol initiated 12-14 days after the presynchronization. Blood was sampled and analyzed for progesterone twice, 12-14 days apart, to determine whether cows had initiated onset of estrous cycles after calving. Cows were scored for body condition in the week after calving, and again at AI, between 69 and 82 days postpartum. Pregnancy was diagnosed at 30+/-3 and 58+/-3 days after AI. Farm influenced all reproductive outcomes evaluated. More (P<0.0001) multiparous than primiparous cows had initiated estrous cycles. Onset of estrous cycles was also influenced (P<0.01) by BCS at calving and at AI, BCS change, season, and milk yield. More (P<0.001) cows that had initiated estrous cycles than anestrous cows were pregnant at 30 and 58 days after AI, but anestrus did not affect pregnancy loss. Conception rates were also influenced (P<0.01) by parity, BCS at calving and AI, BCS change, and season; however, milk yield and insemination protocol were not associated with conception rates at 30 and 58 days after AI. Factors that reduced conception rate on day 30 after AI also increased pregnancy loss between 30 and 58 days of gestation. Improving BCS at calving and AI, minimizing losses of BCS after calving, and hastening onset of estrous cycles early postpartum are all expected to increase conception because of enhanced embryonic survival.

Follicle deviation and ovulatory capacity in Bos indicus heifers

The objectives of Experiment 1 were to determine the interval from ovulation to deviation, and diameter of the dominant follicle (DF) and largest subordinate follicle (SF) at deviation in Nelore (Bos indicus) heifers by two methods (observed and calculated). Heifers (n = 12) were examined ultrasonographically every 12 h from ovulation (Day 0) to Day 5. The time of deviation and diameter of the DF and largest SF at deviation did not differ (P>0.05) between observed and calculated methods. Overall, deviation occurred 2.5+/-0.2 d (mean +/- S.E.M.) after ovulation, and diameters for DF and largest SF at deviation were 6.2+/-0.2 and 5.9 +/- 0.2 mm, respectively. Experiment 2 was designed to determine the size at which the DF acquires ovulatory capacity in B. indicus heifers. Twenty-nine heifers were monitored every 24 h by ultrasonography, from ovulation until the DF reached diameters of 7.0-8.4 mm (n=9), 8.5-10.0 mm (n=10), or >10.0 mm (n=10). At that time, heifers were treated with 25 mg of pLH and monitored by ultrasonography every 12 h for 48 h. Ovulation occurred in 3 of 9, 8 of 10, and 9 of 10 heifers, respectively (P<0.05). In summary, there was no significant difference between observed and calculated methods of determining the beginning of follicle deviation. Deviation occurred 2.5 d after ovulation when the DF reached 6.2 mm, and ovulatory capacity was acquired by DF as small as 7.0 mm.

Ovarian follicle diameter at timed insemination and estrous response influence likelihood of ovulation and pregnancy after estrous synchronization with progesterone or progestin-based protocols in suckled Bos indicus cows.

The objectives of the present study were to evaluate factors associated with estrous synchronization responses and pregnancy per insemination (P/AI) in Bos indicus beef cows submitted to progesterone-based fixed-time artificial insemination (FTAI) protocols. A total of 2388 cows (1869 Nellore and 519 crossbred NellorexAngus) from 10 commercial farms were evaluated to determine the relationships among breed, body condition score (BCS) on the first day of the FTAI protocol, the occurrence of estrus between progesterone device removal and FTAI, and diameter of largest ovarian follicle (LF) at FTAI on estrous synchronization responses and P/AI. Cows (n=412 primiparous; 1976 multiparous) received an intravaginal device containing progesterone or an ear implant containing norgestomet (a progestin), and an injection of estradiol at the beginning of the estrous synchronization protocol. Body condition was scored using a 1-5 scale on the first day of the FTAI protocol and at 30-60 days postpartum. Females received 300IU of equine chorionic gonadotropin (eCG) and PGF(2alpha) on the day the progesterone device/implant was removed and were inseminated 48-60h later. At insemination, cows (n=2388) were submitted to an ultrasonographic exam to determine the diameter of the LF. Follicles were classified into four categories based on mean and standard deviation (SD) of the LF (LF1=two SD below the mean; LF2=mean minus one SD; LF3=mean plus one SD; LF4=two SD above the mean). Ovulation rate was determined in a subset of cows (n=813) by three consecutive ultrasonographic exams: (1) at time of progesterone device/implant removal, (2) at time of FTAI and (3) 48h after FTAI. Ovulation was defined as the disappearance of a large follicle (>or=8.0mm) that was previously recorded. Estrus was determined in a subset of the cows (n=445) by the activation of a detection of estrous patch placed on the tail head on the day of progesterone device/implant removal. Pregnancy was diagnosed 30 days after FTAI. Pregnancy was influenced (P=0.001) by follicle diameter [LF1=27.5% (81/295), LF2=46.6% (328/705), LF3=57.9% (647/1118), LF4=63.3% (171/270)] and the occurrence of estrus [estrus=67.7% (174/257) and no estrus=36.2% (68/188)]. Follicle diameter at FTAI influenced ovulation rate [LF1=42.5% (34/80), LF2=73.9% (161/218), LF3=95.8% (407/425), LF4=97.8% (88/90)], the occurrence of estrus [LF1=54.8% (51/93), LF2=33.6% (43/128), LF3=68.9% (126/183), LF4=90.2% (37/41)] and P/AI among cows that had ovulations [LF1=32.4% (11/34), LF2=50.3% (81/161), LF3=60.0% (244/407), LF4=68.2% (60/88)]. Improving estrous responses between progesterone device withdrawal and FTAI and increasing the diameter of the LF at FTAI may be important aspects to achieve improved estrous synchronization responses and P/AI following progesterone/progestin and estradiol based FTAI protocols in suckled Bos indicus cows.

Effect of intrauterine infusion of ceftiofur on uterine health and fertility in dairy cows.

Objectives were to determine the effects of intrauterine (i.u.) infusion of ceftiofur hydrochloride on uterine health and fertility of dairy cows already receiving PGF(2alpha) for estrous synchronization. Holstein cows at 44 +/- 3 d in milk (DIM) were blocked by parity and diagnosis of metritis in the first 14 DIM and of other illnesses and, within each block, randomly assigned to 1 of 2 treatments: a single i.u. infusion of 125 mg of ceftiofur hydrochloride at 44 +/- 3 DIM (ceftiofur, n = 396), or no i.u. infusion (control, n = 416). All cows received 25 mg of PGF(2alpha) at 37 +/- 3 and 51 +/- 3 DIM as part of an estrous synchronization protocol. A subset of 547 cows was evaluated for clinical endometritis immediately before treatment, and 202 cows had an aseptic uterine sample collected before the injection of PGF(2alpha) at 51 +/- 3 DIM for bacteriology and diagnosis of subclinical endometritis (> or =5% neutrophils). Pregnancy on d 38 +/- 3 and 180 +/- 7 after the first artificial insemination, pregnancy loss, and interval from calving to pregnancy in the first 300 DIM were evaluated. The proportions of cows diagnosed with clinical endometritis before treatment were similar between ceftiofur and control treatments. Intrauterine infusion with ceftiofur did not influence prevalence of subclinical endometritis and positive uterine culture 7 d after treatment; however, it reduced the prevalence of positive uterine culture in cows with clinical endometritis (29.0 vs. 51.4%) and reduced the overall prevalence of Arcanobacterium pyogenes (1.0 vs. 7.6%) at 51 +/- 3 DIM. Cows with clinical endometritis had increased prevalence of A. pyogenes (10.3 vs. 1.5%), Escherichia coli (5.9 vs. 0.75%), and overall positive uterine culture (41.2 vs. 22.4%); however, cows with subclinical endometritis only had an increased prevalence of A. pyogenes (10.2 vs. 1.5%). Ceftiofur did not affect pregnancy per artificial insemination in all cows or in cows previously diagnosed with metritis or clinical endometritis. Interval to pregnancy was similar for control and ceftiofur cows. Intrauterine infusion of ceftiofur hydrochloride reduced the prevalence of uterine infection in cows with clinical endometritis, and the prevalence of A. pyogenes, but did not affect the prevalence of subclinical endometritis or fertility of dairy cows already receiving PGF(2alpha).

Equine chorionic gonadotropin and gonadotropin-releasing hormone enhance fertility in a norgestomet-based, timed artificial insemination protocol in suckled Nelore (Bos indicus) cows

Two experiments were conducted to investigate the effects of equine chorionic gonadotropin (eCG) at progestin removal and gonadotropin-releasing hormone (GnRH) at timed artificial insemination (TAI) on ovarian follicular dynamics (Experiment 1) and pregnancy rates (Experiment 2) in suckled Nelore (Bos indicus) cows. Both experiments were 2x2 factorials (eCG or No eCG, and GnRH or No GnRH), with identical treatments. In Experiment 1, 50 anestrous cows, 134.5+/-2.3 d postpartum, received a 3mg norgestomet ear implant sc, plus 3mg norgestomet and 5mg estradiol valerate im on Day 0. The implant was removed on Day 9, with TAI 54 h later. Cows received 400 IU eCG or no further treatment on Day 9 and GnRH (100 microg gonadorelin) or no further treatment at TAI. Treatment with eCG increased the growth rate of the largest follicle from Days 9 to 11 (means+/-SEM, 1.53+/-0.1 vs. 0.48+/-0.1mm/d; P<0.0001), its diameter on Day 11 (11.4+/-0.6 vs. 9.3+/-0.7 mm; P=0.03), as well as ovulation rate (80.8% vs. 50.0%, P=0.02), whereas GnRH improved the synchrony of ovulation (72.0+/-1.1 vs. 71.1+/-2.0 h). In Experiment 2 (n=599 cows, 40 to 120 d postpartum), pregnancy rates differed (P=0.004) among groups (27.6%, 40.1%, 47.7%, and 55.7% for Control, GnRH, eCG, and eCG+GnRH groups). Both eCG and GnRH improved pregnancy rates (51.7% vs. 33.8%, P=0.002; and 48.0% vs 37.6%, P=0.02, respectively), although their effects were not additive (no significant interaction). In conclusion, eCG at norgestomet implant removal increased the growth rate of the largest follicle (LF) from implant removal to TAI, the diameter of the LF at TAI, and rates of ovulation and pregnancy rates. Furthermore, GnRH at TAI improved the synchrony of ovulations and pregnancy rates in postpartum Nelore cows treated with a norgestomet-based TAI protocol.

Equine chorionic gonadotropin improves the efficacy of a progestin-based fixed-time artificial insemination protocol in Nelore (Bos indicus) heifers.

A total of 177 Nelore heifers were examined by ultrasonography to determine the presence or absence of a corpus luteum (CL) and received a 3mg norgestomet ear implant plus 2mg of estradiol benzoate i.m. On Day 8, implants were removed and 150 microg of d-cloprostenol i.m. was administered. At the time of norgestomet implant removal, heifers with or without CL at the time of initiating treatment were assigned equally and by replicate to be treated with 0IU (n=87) or 400IU (n=90) eCG i.m. All heifers received 1mg of EB i.m. on Day 9 and were submitted to fixed-time artificial insemination (FTAI) 30-34h later. The addition of eCG increased the diameter of the largest follicle (LF) at FTAI (10.6+/-0.2mm vs. 9.5+/-0.2mm; P=0.003; mean+/-SEM), the final growth rate of the LF (1.14+/-0.1mm/day vs. 0.64+/-0.1mm/day; P=0.0009), ovulation rate [94.4% (85/90) vs. 73.6% (64/87); P=0.0006], the diameter of the CL at Day 15 (15.5+/-0.3mm vs. 13.8+/-0.3mm; P=0.0002), serum concentrations of progesterone 5 days after FTAI (6.6+/-1.0 ng/ml vs. 3.6+/-0.7ng/ml; P=0.0009), and pregnancy per AI [P/AI; 50.0% (45/90) vs. 36.8% (32/87); P=0.04]. The absence of a CL at the beginning of the treatment negatively influenced the P/AI [30.2% (16/53) vs. 49.2% (61/124); P=0.01]. Therefore, the presence of a CL (and/or onset of puberty) must be considered in setting up FTAI programs in heifers. In addition, eCG may be an important tool for the enhancement of follicular growth, ovulation, size and function of the subsequent CL, and pregnancy rates in progestin-based FTAI protocols in Bos indicus heifers.

Importance of estrus on pregnancy per insemination in suckled Bos indicus cows submitted to estradiol/progesterone-based timed insemination protocols.

The objective was to evaluate the effect of estrus occurrence (based on removal of tail-head marks) on ovarian responses and pregnancy per AI (P/AI; 30 d after AI) in suckled Bos indicus beef cows submitted to timed AI (TAI) protocols. Cows received an intravaginal device containing 1.0 g progesterone, and 2.0 mg estradiol benzoate im; 8 d later, the intravaginal device was removed, and they were given PGF(2α) (0.25 mg of cloprostenol sodium) and 300 IU of eCG, with TAI 48 to 52 h later. In Experiment 1, cows were assigned to receive one of three treatments: 1 mg of estradiol cypionate (ECP) im at progesterone (P4) device removal (N = 178); 10 μg of GnRH im at TAI (N = 190); or both treatments (N = 172). In cows given estradiol (ECP or ECP + GnRH), more displayed estrus (P = 0.002) and became pregnant (P < 0.0001) compared with those receiving only GnRH. In Experiment 2, the effect of the occurrence of estrus on ovarian responses was evaluated in cows (N = 53) synchronized using ECP at device removal. Cows that displayed estrus had a greater diameter of the largest follicle (LF) at device removal (P < 0.0001), a greater diameter at TAI (P < 0.0001), a greater ovulation rate (P = 0.02), a larger CL (P = 0.02), and a greater P4 concentration (P < 0.0001) than cows that did not display estrus. In Experiment 3, the effect of GnRH treatment on P/AI at TAI was evaluated in cows that received ECP at device removal, and either displayed, or did not display, estrus (N = 726). There was no estrus by GnRH interaction (P = 0.22); the P/AI was greater (P < 0.0001) in cows that displayed estrus (61.9%) than cows that did not display estrus (41.4%). However, GnRH did not improve (P = 0.81) P/AI (GnRH = 53.7% vs. no GnRH = 52.6%). In conclusion, exogenous estradiol at device removal increased both the proportion of suckled Bos indicus cows that displayed estrus and P/AI. Cows that displayed estrus had better ovarian responses (i.e., larger follicles at TAI, a greater ovulation rate, larger CL, and greater P4 concentrations) following an estradiol/P4-based synchronization protocol. Although occurrence of estrus improved pregnancy outcomes, GnRH at TAI did not improve P/AI in suckled Bos indicus cows treated with ECP, regardless of estrus occurrence.

Timing of insemination and fertility in dairy and beef cattle receiving timed artificial insemination using sex-sorted sperm

The objective was to evaluate the effects of timing of insemination and type of semen in cattle subjected to timed artificial insemination (TAI). In Experiment 1, 420 cyclic Jersey heifers were bred at either 54 or 60 h after P4-device removal, using either sex-sorted (2.1 × 10(6) sperm/straw) or non-sorted sperm (20 × 10(6) sperm/straw) from three sires (2 × 2 factorial design). There was an interaction (P = 0.06) between time of AI and type of semen on pregnancy per AI (P/AI, at 30 to 42 d after TAI); it was greater when sex-sorted sperm (P < 0.01) was used at 60 h (31.4%; 32/102) than at 54 h (16.2%; 17/105). In contrast, altering the timing of AI did not affect conception results with non-sorted sperm (54 h = 50.5%; 51/101 versus 60 h = 51.8%; 58/112; P = 0.95). There was an effect of sire (P < 0.01) on P/AI, but no interaction between sire and time of AI (P = 0.88). In Experiment 2, 389 suckled Bos indicus beef cows were enrolled in the same treatment groups used in Experiment 1. Sex-sorted sperm resulted in lower P/AI (41.8%; 82/196; P = 0.05) than non-sorted sperm (51.8%; 100/193). In addition, there was a tendency for greater P/AI (P = 0.11) when TAI was performed 60 h (50.8%; 99/195) versus 54 h (42.8%; 83/194) after removing the progestin implant. In Experiment 3, 339 suckled B. indicus cows were randomly assigned to receive TAI with sex-sorted sperm at 36, 48, or 60 h after P4 device removal. Ultrasonographic examinations were performed twice daily in all cows to confirm ovulation. On average, ovulation occurred 71.8 ± 7.8 h after P4 removal, and greater P/AI was achieved when insemination was performed closer to ovulation. The P/AI was greatest (37.9%) for TAI performed between 0 and 12 h before ovulation, whereas P/AI was significantly less for TAI performed between 12.1 and 24 h (19.4%) or >24 h (5.8%) before ovulation. In conclusion, sex-sorted sperm resulted in a lesser P/AI than non-sorted sperm following TAI. However, improvements in P/AI with delayed time of AI were possible (Experiments 1 and 3), and seemed achievable when breeding at 60 h following progestin implant removal, compared to the standard 54 h normally used in TAI protocols.

Effects of feeding rumen-protected choline on incidence of diseases and reproduction of dairy cows.

The objectives of this study were to determine the effects of feeding rumen-protected choline (RPC) to dairy cows on the incidence of disease in early lactation, and on their fertility as measured by return to ovarian cyclicity and pregnancies per insemination (P/AI). In the first experiment, 369 cows were fed 15 g/day of RPC between 25 days pre-calving and 80 days post calving. In the second experiment, 578 primigravid cows were fed 15 g/day of RPC for the last 21 days of gestation only. In both experiments, P/AI were evaluated for the first and second inseminations, and health disorders were monitored daily until 90 days after calving. Return to cyclicity was evaluated in experiment 1 only, on days 51 and 61 post calving. In the first experiment, feeding RPC reduced the incidence of clinical ketosis, mastitis, and morbidity, and the number of cases of mastitis per cow. Feeding RPC did not influence cyclicity and P/AI. In the second experiment, cows fed RPC tended to have greater morbidity than controls because of an increased incidence of metritis and fever, although the incidence of retained fetal membranes was lower. The P/AI for first and second inseminations were similar between treatments. Supplementing the diets of dairy cattle with RPC from before until after calving improved the health of early lactation cows. However, in primigravid cows feeding RPC before calving only had mixed effects on health. Feeding RPC had no beneficial effects on reproduction in either experiment.

Manipulation of Follicle Development to Ensure Optimal Oocyte Quality and Conception Rates in Cattle

Over the last several decades, a number of therapies have been developed that manipulate ovarian follicle growth to improve oocyte quality and conception rates in cattle. Various strategies have been proposed to improve the responses to reproductive biotechnologies following timed artificial insemination (TAI), superovulation (SOV) or ovum pickup (OPU) programmes. During TAI protocols, final follicular growth and size of the ovulatory follicle are key factors that may significantly influence oocyte quality, ovulation, the uterine environment and consequently pregnancy outcomes. Progesterone concentrations during SOV protocols influence follicular growth, oocyte quality and embryo quality; therefore, several adjustments to SOV protocols have been proposed depending on the animal category and breed. In addition, the success of in vitro embryo production is directly related to the number and quality of cumulus oocyte complexes harvested by OPU. Control of follicle development has a significant impact on the OPU outcome. This article discusses a number of key points related to the manipulation of ovarian follicular growth to maximize oocyte quality and improve conception rates following TAI and embryo transfer of in vivo- and in vitro-derived embryos in cattle.