Effects of additional gonadotropin-releasing hormone and prostaglandin F2α treatment to an estradiol/progesterone-based embryo transfer protocol for recipient lactating dairy cows.

This study was designed to evaluate whether the utilization of a second PGF2α treatment at the end of an estradiol/progesterone (E2/P4)-based protocol with or without GnRH at the beginning of the protocol would improve pregnancy rates of lactating Holstein cows assigned to timed embryo transfer. A total of 501 lactating Holstein cows in 5 farms were enrolled in the experiment. Within farm, cows were blocked by parity and, within block, were assigned randomly to (1) insertion of an intravaginal P4 device (controlled internal drug-releasing device; CIDR) and estradiol benzoate on d -11, PGF2α on d -4, CIDR withdrawal and an injection of estradiol cypionate on d -2, and timed embryo transfer on d 7 (1-PGF; n = 164); (2) the same treatments as 1-PGF, but with PGF2α administered on d -4 and -2 (2-PGF; n = 171); and (3) 2-PGF with the addition of a GnRH treatment on d -11 (GnRH+2-PGF; n = 166). Ovaries were scanned by transrectal ultrasonography on d -11, -4, and 7, and blood samples were collected on d -11, -4, 0, and 7 for P4 determination. Treatment comparisons were performed using contrasts. The proportion of cows with a new corpus luteum on d -4 was greater in GnRH+2-PGF cows. Cows in 1-PGF had a greater P4 concentration on d 0 but lesser P4 on d 7 compared with cows in the other groups. Cows assigned to receive 2-PGF (2-PGF and GnRH+2-PGF) had greater estrus expression, and a greater proportion of cows ovulated to estradiol cypionate. No further contrast effects were observed for follicle diameter, double ovulation rate, pregnancy per embryo transfer (P/ET) on d 32 and 60, or pregnancy loss. As P4 concentration on d -4 increased, P/ET on d 60 tended to increase. Cows with P4 ≥3.66 ng/mL on d -4 had greater P/ET on d 32 and 60 than those with P4 below that threshold. Regardless of treatment, cows with P4 concentration ≥3.66 ng/mL also had a greater pregnancy per synchronized protocol (P/SP) on d 60. Also, a P4 concentration on d -4 (low or high) × follicle diameter (continuous) interaction tendency was observed when evaluating P/ET. Although P/ET did not differ among cows with different follicles sizes with reduced P4 concentration on d -4 (<3.66 ng/mL), it increased in cows with larger follicles exposed to increased P4 concentration (≥3.66 ng/mL). When P4 on d 0 was evaluated, P/ET on d 32 and 60 was greater for cows with low (≤0.09 ng/mL) versus high (>0.21 ng/mL) P4; as P4 concentration on d 0 increased, P/ET linearly decreased. In summary, cows with increased P4 concentrations during growth of the ovulatory follicular wave had improved P/ET. Administering a second PGF2α dose reduced P4 concentration on d 0 and increased ovulatory response to the protocol, but no benefits were observed on P/ET or P/SP.

Somatic cell count and type of intramammary infection impacts fertility from in vitro produced embryo transfer.

The objective of this study was to assess the impact of mastitis-causing bacteria and somatic cell count (SCC) on pregnancy per embryo transfer (P/ET) in Holstein-Gir crossbred (Girolando) lactating dairy cows. Cows (n = 1397) were subjected to a timed-embryo transfer protocol. Milk samples were collected two days before embryo transfer for SCC and bacteriological culture analyses. Pregnancy diagnosis was performed on days 31 and 66 after timed-embryo transfer. The animals were grouped according to the National Mastitis Council recommendations: Gram-positive environmental (EV+), Gram-negative environmental (EV-), Gram-positive contagious (C+), coagulase-negative staphylococci (CNS) and control (no bacterial growth). Additional analysis was made by categorizing bacteria based on degree of pathogenicity (Major or Minor). Bacterial growth reduced P/ET (P < .01) at both 31 and 66 days of gestation. The P/ET was lower (P < .05) at 31 days in EV- (30.1%) and EV+ (29.9%) groups and tended (P = .09) to be lower in the C+ group (36.6%) than the control group (44.0%). The P/ET from the Major group at 31 days of gestation was lower (P = .03) compared with the Minor and control groups (32.1 vs 41.1 vs 43.2%, respectively). Cows with SCC > 400,000 cells/mL had lower P/ET (P < .01) than animals with SCC < 200,000 cells/mL at both 31 (30.4% vs 40.8%) and 66 days (24.7% vs 32.2%) of gestation. Pregnancy loss was not different between bacterial isolates and SCC categories. Elevated SCC significantly reduced P/ET, whereas environmental agents and those with Major pathogenicity yielded the greatest reduction in P/ET.

Effect of adding a gonadotropin-releasing-hormone treatment at the beginning and a second prostaglandin F2α treatment at the end of an estradiol-based protocol for timed artificial insemination in lactating dairy cows during cool or hot seasons of the year.

Our hypothesis was that fertility could be increased in a timed artificial insemination (TAI) protocol based on estradiol (E2) and progesterone (P4) by combining GnRH with E2-benzoate at the start of the protocol to increase circulating P4 during preovulatory follicle development and by using 2 prostaglandin F2α (PGF) treatments at the end to decrease P4 near TAI. Lactating Holstein cows (n=1,808) were randomly assigned during the cool or hot season of the year to receive TAI (d 0) following 1 of 3 treatments: (1) control: controlled internal drug-release insert + 2mg of E2-benzoate on d -11, PGF on d -4, controlled internal drug-release insert withdrawal + 1.0mg of E2-cypionate on d -2, and TAI on d 0; (2) 2PGF: identical to control protocol with addition of a second PGF treatment on d -2; (3) GnRH: identical to 2PGF protocol with addition of a 100-µg GnRH treatment on d -11. Pregnancy diagnoses were performed on d 32 and 60 after TAI. Season had major effects on many reproductive measures, with cool season greater than hot season in percentage of cows with corpus luteum (CL) at PGF (62.9 vs. 56.2%), ovulatory follicle diameter (15.7 vs. 14.8mm), expression of estrus (86.7 vs. 79.9%), ovulation following the protocol (89.7 vs. 84.3%), and pregnancies per artificial insemination (P/AI; 45.4 vs. 21.4%). The GnRH protocol increased percentage of cows with CL (control=56.9%; 2PGF=55.8%; GnRH=70.5%) and P4 at PGF (control=3.28±0.22; 2PGF=3.35±0.22; GnRH=3.70±0.21ng/mL), compared with control and 2PGF protocols. The GnRH protocol increased P/AI at the pregnancy diagnosis at 32d [37.3% (219/595)] and 60d [31% (179/595)] after TAI, compared with control [30.0% (177/604); 25.1% (145/604)], with intermediate results with 2PGF protocol [33.2% (196/609); 28.0% (164/609)]. The positive effects of GnRH treatment on P/AI were only detected during the cool season (GnRH=50.9%; 2PGF=44.2%; control=41.0%) and not during the hot season. In addition, the effect of GnRH was only observed in cows with low P4 (<3ng/mL) at the start of the protocol and not in cows that began the protocol with high P4. Furthermore, presence of CL at PGF interacted with follicle diameter such that cows with a CL at PGF had greater P/AI if they ovulated larger rather than smaller follicles near TAI. Thus, fertility to TAI can be improved by inducing ovulation at the beginning of an E2/P4-based protocol using GnRH treatment, particularly during the cool season of the year and in cows with low P4 at the start of the protocol.

Targeted progesterone supplementation improves fertility in lactating dairy cows without a corpus luteum at the initiation of the timed artificial insemination protocol.

The objectives of this study were to determine the effect of supplemental progesterone on fertility in lactating dairy cows lacking a corpus luteum (CL) at the initiation of the timed artificial insemination (AI) program. Holstein cows were subjected to the 5-d timed AI program (d -8 GnRH, d -3 and -2 PGF2α, d 0 GnRH and AI). Cows had their ovaries scanned by ultrasonography on d -8 and those bearing a CL were considered to be in diestrus (DI; n=946). Cows that lacked a CL on d -8 were assigned to remain as untreated control (CON; n=234) or receive 2 controlled internal drug release (CIDR) inserts containing progesterone (2CIDR; n=218) from d -8 to -3, as a single insert has been proven insufficient to modulate fertility in cows without CL. Blood was analyzed for progesterone and estradiol concentrations. Pregnancy was diagnosed on d 34 and 62 after AI. Progesterone concentrations during the timed AI program were lowest for CON, intermediate for 2CIDR, and highest for DI. Supplementation increased progesterone concentrations between d -7 and -3 compared with CON (2.65 vs. 0.51 ng/mL). Ovulation to the first GnRH was not affected by treatment. However, a greater proportion of CON and 2CIDR cows had a new CL on d -3 compared with DI cows (66.7 vs. 61.9 vs. 52.0%). In cows with a new CL, the diameter of the ovulatory follicle was larger for CON than 2CIDR, and intermediate for DI (18.7 vs. 16.5 vs. 17.7 mm). Concentrations of estradiol on d -3 did not differ among treatments; however, DI cows had greater estradiol concentrations at AI compared with CON or 2CIDR cows. Pregnancy per AI was less for CON compared with 2CIDR or DI on d 32 (30.8 vs. 46.8 vs. 49.9%) and 64 (28.6 vs. 43.7 vs. 47.3%), indicating that supplementation with progesterone reestablished fertility in cows lacking a CL on d -8. A greater proportion of nonpregnant CON cows had a short reinsemination interval compared with 2CIDR or DI (11.1 vs. 3.5 vs. 5.7%). Treatment did not affect pregnancy loss between d 34 and 62 of gestation. A single ultrasound exam was effective in identifying a low-fertility cohort of cows based on the absence of CL at the first GnRH injection of the timed AI protocol. Progesterone supplementation with 2 CIDR inserts increased progesterone in plasma to 2.65 ng/mL and restored fertility in lactating dairy cows lacking a CL at the initiation of the timed AI program similar to that of cows in diestrus.