The effectiveness of low-dose of eCG in timed AI Nelore (Bos indicus) heifers.

This study evaluated the efficacy of the administration of different doses of equine chorionic gonadotropin (eCG; 0 IU, 200 IU, or 300 IU) at the time of the progesterone device removal in 2-year-old Nelore (Bos indicus) heifers synchronized for fixed-timed artificial insemination (FTAI). On day 0 (D0), a total of 398 heifers received 2 mg of oestradiol benzoate i.m., 0.53 mg of cloprostenol i.m., and an eight-day previously used (second use) intravaginal device containing 1 g of progesterone (P4). Eight days later (D8), simultaneous with the P4 device removal, 0.5 mg of oestradiol cypionate i.m. and 0.53 mg of cloprostenol i.m. were administered. At the same time, heifers were randomly assigned to receive one of the following treatments: G-0 IU (n = 141; no eCG treatment), G-200 IU (n = 132; treated with 200 IU of eCG), and G-300 IU (n = 125; treated with 300 IU of eCG). FTAI was performed 48 h after the P4 device removal (D10). Ultrasonographic evaluations were performed at D0, D10, and D17. Heifers were scanned to measure the size of the largest follicle (LF), the presence, number, and size of the corpus luteum (CL), and the ovulation rate. Subsequently, at D40, the heifers underwent scanning to determine the pregnancy rate and identify any twin pregnancies. Additionally, at D70, scans were performed to assess pregnancy loss (PG). Data were analysed by orthogonal contrasts [C1 (eCG effect): control x (200 IU + 300 IU) and C2 (eCG dose effect): 200 IU × 300 IU]. On D0, CL presence was similar between the groups [G-0 IU = 65.2% (92/141), G-200 IU = 55.3% (73/132), and G-300 IU = 63.2% (79/125); p = .16]. No interactions between the presence of CL on D0 and eCG treatment were found for any of the variables (p > .05). The diameter of the LF at FTAI (D10) was not influenced by eCG treatment (p = .22) or eCG dose (p = .18). However, treatment with eCG increased the diameter of the CL at D17 (G-0 IU = 15.7 ± 0.3 mmb , G-200 IU = 16.6 ± 0.2 mma , and G-300 IU = 16.6 ± 0.3 mma ; p = .001), regardless of the dose used (p = .94). The ovulation rate was higher in heifers treated with eCG [G-0 IU = 79.4%b (112/141), G-200 IU = 90.2%a (119/132), and G-300 IU = 93.6%a (117/125); p = .002], but there was no effect of eCG dose (p = .36). Pregnancy per AI (P/AI) on D40 [G-0 IU = 32.6%b (46/141), G-200 IU = 42.4%a (56/132), and G-300 IU = 42.4%a (53/125); P = 0.05] and D70 [G-0 IU = 29.1%b (41/141), G-200 IU = 40.9%a (54/132), and G-300 IU = 40.8%a (51/125); p = .02] were higher on heifers that received eCG; however, no dose effect was observed for P/AI on D40 (p = .89) nor D70 (p = .98). Pregnancy loss between D40 and D70 tended to reduce (p = .07) in eCG-treated heifers without dose effect (p = .91). Heifers with CL at D0 presented a greater follicle diameter (LF) on D10 (With CL = 11.2 ± 0.2 mm and Without CL = 10.2 ± 0.2 mm; p = .05), CL diameter on D17 (With CL = 15.8 ± 0.03 mm and Without CL = 11.8 ± 0.6 mm; p = .01), and ovulation rate [With CL = 95.5% (233/244) and Without CL = 74.7% (115/154); p = .01]. However, no difference in pregnancy rate at D40 (p = .52) and D70 (p = .84) was found. In conclusion, eCG treatment increases ovulation and pregnancy rates of heifers submitted to a FTAI protocol. Furthermore, eCG treatment increases the diameter of the CL after FTAI and reduces pregnancy losses. No dose effect was observed, suggesting Nelore (Bos indicus) heifers respond to 200 IU of eCG treatment for FTAI.

Use of injectable progesterone at the beginning of the TAI protocol is not necessary in super-early resynchronization started 14 days after artificial insemination in Bos indicus beef heifers.

In a protocol for the resynchronization of ovulation starting 14 days (D14) after timed artificial insemination, the effects of short-action injectable progesterone (P4i) administration and the length of treatment with an intravaginal progesterone (P4) device on follicular dynamics, the conception rate (P/AI) and the percentage of false positives were evaluated in 1065 Nelore heifers previously timed-inseminated. On D14, P4 devices were inserted, and heifers allocated, based on a 2 × 2 factorial design, to receive (P4i) or not 50 mg of P4i (NoP4i), and to remove the P4 device after 6 (6Day) or 8 days (8Day). At the time of P4 device removal (D20 and D22), non-pregnancy diagnosis was performed using vascularization of the corpus luteum (VCL) evaluated. At this time, non-pregnant heifers received 150 µg of D-cloprostenol, 0.6 mg of estradiol cypionate and 300IU of eCG. TAI was performed 48 h after P4 device removal. For these heifers, ultrasound examinations were performed at P4 device removal and at TAI to evaluate follicular dynamics and at 30 days after TAI to evaluate the P/AI. Pregnant heifers based on VCL were evaluated using B-mode ultrasonography 10 days after Doppler ultrasound to evaluate the percentage of false positives. Statistical analyses were performed using the GLIMMIX procedure of SAS. There were no interaction effects between P4i and duration of treatment with a P4 device. The P/AI was diagnosed by Doppler, 1st TAI, 2nd TAI, total and percentage of false positives did not differ between heifers receiving or not P4i. Similarly, duration of treatment with a P4 device did not influence the P/AI by Doppler, 1st TAI, 2nd TAI and total. However, the percentage of false positives diagnoses was higher in 6Day heifers (P = 0.01). The diameter of largest follicle at P4 device removal was greater in the 8Day heifers (P = 0.001), and at TAI was higher in the P4i-treated heifers (P = 0.03). Additionally, the percentage of false positives diagnoses was lower in heifers that ovulated to the 1st TAI protocol (P = 0.001). In conclusion, for resynchronization 14 days after TAI, it is not necessary to inject P4i at the beginning of the protocol. In addition, P4 device removal after 6 instead of 8 days increases the percentage of false positives because of the earlier diagnosis (20 days after TAI), but does not interfere in P/AI of resynchronization protocol. Furthermore, the percentage of false positives is higher in heifers that did not ovulate to 1st TAI protocol.

Temporal evaluation of follicular dynamics and endocrine patterns of Holstein (Bos taurus), Gir (Bos indicus), and Murrah (Bubalus bubalis) heifers kept under the same nutritional, management and environmental conditions.

The objective of the study was to simultaneously compare ovarian follicular dynamics and endocrine parameters of taurine (Holstein; n = 14), zebuine (Gir; n = 5), and bubaline (Murrah; n = 15) heifers kept under the same environmental, nutritional and management conditions. Heifers were synchronized with two PGF treatments 14 days apart. Ovaries of cyclic heifers were scanned daily during two consecutive ovulations and blood samples were collected every 24 h from each animal. No significant difference was found for length of interovulatory interval, however, zebuine heifers presented a greater number of follicular waves, number of antral follicles on day of ovulation, and higher insulin concentration than the other two breeds. Taurine heifers had highest maximal diameter of first wave dominant and ovulatory follicles and CL volume. Taurine and bubaline heifer's dominant follicle of first wave had longer static and regression phases than zebuine heifers. Bubaline heifers presented overall lowest progesterone concentrations and CL volume, but higher IGF1 levels. No difference was observed between taurine and zebuine heifers regarding IGF1 concentration. Despite higher CL volume found in taurine heifers, no difference in mean progesterone concentration was observed between them and zebuine heifers. Insulin and IGF1 concentrations were greater in follicular phase than in luteal phase when breed was not evaluated. After evaluating the three breeds simultaneously, at the same nutritional and management status it is possible to conclude that each genetic group has a specific follicular development and endocrinology of the estrous cycle.

Assessment of the main pathogens associated with clinical and subclinical endometritis in cows by culture and MALDI-TOF mass spectrometry identification.

Clinical endometritis (CE) and subclinical endometritis (SCE) are diseases that affect dairy cows during the puerperium, causing negative effects on the animals' milk production and fertility. The objective of this study was to assess the main bacteria related to cases of CE and SCE from uterine samples of dairy cows in Brazilian herds. Selective and differential media were used for isolation of aerobic and anaerobic bacteria and further MALDI-TOF mass spectrometry (MS) identification. A total of 279 lactating dairy cows with 28 to 33 d in milk from 6 commercial farms were evaluated. Initially, cows were classified in 3 groups: cytologic healthy cows (n = 161), cows with CE (n = 83), and cows with SCE (n = 35). Healthy animals presented 97 species, followed by the CE group with 53 identified species, and SCE cows presented only 21 bacterial species. We found a significantly higher isolation rate of Trueperella pyogenes in CE (26.5%) cows compared with healthy and SCE cows. Some anaerobic species were exclusively isolated from the CE group, even though they presented lower frequency. Interestingly, 18.1% of samples from CE cows and 40% of SCE cows were negative to bacterial isolation. Despite the use of culture-dependent methods instead of molecular methods, the present study enabled the identification of a complex community of 127 different species from 48 genera, composed of aerobic and anaerobic bacterial species among the 3 different animal groups. The method of sample collection, culture, and identification by MALDI-TOF MS were essential for the success of the analyses.

Use of cooled buffalo semen as a strategy to increase conception rates in fixed-time artificial insemination programs during unfavorable reproductive periods / [Uso de sêmen resfriado de búfalo como estratégia para aumentar as taxas de concepção em programas de inseminação artificial em tempo fixo, durante períodos reprodutivos desfavoráveis]

The objective of this study was to compare the reproductive efficiency of dairy buffaloes undergoing fixed-time artificial insemination (FTAI) protocols based on progesterone/estrogen (P4/E2) and eCG during unfavorable breeding season using cooled (CS) and frozen semen (FS). A total of 446 buffaloes (> 40 days postpartum) were randomly distributed into four blocks (years): B1-2014 (n = 143), B2-2015 (n = 34), B3-2016 (n = 90), and B4-2017 (n = 179). Each block was subdivided into two (AI with CS and FS using the same ejaculate of each bull). Thus, the block subdivision was as follows: B1 (CS = 71 and FS = 72); B2 (CS = 18 and FS = 16); B3 (CS = 47 and FS = 43); and B4 (CS = 90 and FS = 89). The ejaculates of eight Murrah bulls collected using an artificial vagina were divided into two aliquots: one aliquot was diluted in Botu-Bov® commercial extender and cooled (BB-CS), and the other was diluted in the same extender and frozen (BB-FS). BB-CS aliquots were cooled at 5 °C/24 h using a refrigerator. BB-FS group aliquots were also cooled, and after equilibrating at 5 °C for 4 h, were placed in a 21-L Styrofoam box, 5 cm above the surface of liquid nitrogen. In the afternoon (A) on D0 (2:00 p.m.) the animals received EB 2.0 mg IM (Estrogin®) and an ear implant (CRESTAR® 3.0 mg P4). At D9 (A), the implant was removed, and the animals received eCG 400 IU IM (Folligon® 5000) + Cloprostenol PGF2α 0.530 mg IM (Sincrocio®). At D10 (A), the animals received EB 1.0 mg IM (Estrogin®), and at D12 (8:00 a.m.), AI was performed. At D42, pregnancy was diagnosed via ultrasonography. Total CRs were 48.2% CS and 34.6% FS for years 2014 to 2017, with a significant difference of 13.7% (P<0.05). In conclusion, cooled semen resulted in higher CR than frozen semen in dairy buffaloes under the P4/E2 and eCG FTAI during the unfavorable reproductive season.(AU)

O objetivo deste estudo foi comparar a eficiência reprodutiva de búfalas leiteiras submetidas a protocolos de inseminação artificial em tempo fixo (IATF) à base de progesterona/estrogênio (P4/E2) e eCG, durante a estação reprodutiva desfavorável, usando-se sêmen resfriado (SR) e congelado (SC) Um total de 446 búfalas (> 40 dias após o parto) foi distribuído aleatoriamente em quatro blocos (anos): B1-2014 (n = 143), B2-2015 (n = 34), B3-2016 (n = 90) e B4-2017 (n = 179). Cada bloco foi subdividido em dois (IA com SR e SC utilizando-se a mesma ejaculação de cada touro). Assim, a subdivisão do bloco foi a seguinte: B1 (SR = 71 e SC = 72); B2 (SR = 18 e SC = 16); B3 (SR = 47 e SC = 43); e B4 (SR = 90 e SC = 89). Os ejaculados de oito touros Murrah coletados com vagina artificial foram divididos em duas alíquotas: uma alíquota diluída em diluente comercial Botu-Bov® e resfriada (BB-SR), e a outra diluída no mesmo diluente e congelada (BB-SC). As alíquotas de BB-SR foram resfriados a 5°C/24h usando-se um refrigerador. As alíquotas do grupo BB-SC também foram resfriadas e, após equilíbrio a 5°C por 4h, foram colocadas em uma caixa de isopor de 21L, 5 cm acima da superfície do nitrogênio líquido. À tarde (A), no D0 (14h), os animais receberam BE 2,0 mg IM (Estrogin®) e um implante auricular (Crestar® 3,0 mg P4). No D9 (A), o implante foi retirado e os animais receberam eCG 400 UI IM (Folligon® 5000) + cloprostenol PGF2α 0,530 mg IM (Sincrocio®). No D10 (A), os animais receberam BE 1,0mg IM (Estrogin®), e, no D12 (8h da manhã), foram realizadas as IAs. No D42, a gestação foi diagnosticada por ultrassonografia. As taxas de concepção (TC) totais foram 48,2% SR e 34,6% SC para os anos de 2014 a 2017, com uma diferença significativa de 13,7% (P<0,05). Em conclusão, o sêmen resfriado resultou em maior TC do que o sêmen congelado em bubalinos leiteiros sob P4/E2 e eCG FTAI durante a estação reprodutiva desfavorável.(AU)

Effects of eCG and FSH in timed artificial insemination treatment regimens on estrous expression and pregnancy rates in primiparous and multiparous Bos indicus cows.

Effects were evaluated in Bos indicus cows of eCG and FSH on follicular growth, estrous expression, and pregnancy per artificial insemination (P/AI) as a result of fixed-time artificial insemination (TAI). In Experiment 1, extent of timing-of-ovulation synchronization among cows was evaluated after imposing an estrogen/progesterone-based treatment regimen. At progesterone device removal (D8), cows were administered: eCG, or FSH or served as untreated Controls. In Experiment 2, percentage of cows P/AI was evaluated when the Experiment 1-treatment regimen was imposed. On D10, all cows were artificially inseminated. In Experiment 3, cows were assigned to two treatment groups (Control and eCG) on D8 to evaluate percentage of cows P/AI and estrous expression. In Experiment 1, follicular dynamics were similar among treatment groups. In Experiment 2, follicular growth was greater (P = 0.0001) with the eCG treatment. There was an interaction of treatment × parity (P = 0.007) on percentage of cows P/AI. There was a greater percentage of primiparous cows P/AI in the eCG-treated than Control and FSH-treated cows. There was a greater percentage of eCG-treated multiparous cows pregnant as a result of TAI than Control cows. There was an interaction of treatment × parity (P = 0.005) on P/AI in Experiment 3, in which the eCG effect was more pronounced in primiparous cows. Treatment with FSH, therefore, was not as effective as eCG in stimulation of follicular growth or enhancing percentage of cows pregnant as a result of TAI. Physiological effects of eCG, however, were also more evident in primiparous cows.

Optimization of a 5-day fixed-time embryo transfer (FTET) protocol in heifers I. Manipulation of circulating progesterone through reutilization of intravaginal progesterone devices during FTET.

The objectives of the present study were to: 1) compare the reproductive efficiency of embryo transfer (ET) recipients after synchronization of estrus or a 5-day synchronization of ovulation protocol for fixed time ET (FTET), and 2) determine the effect of reutilization of intravaginal P4 devices (CIDRs), up to four times, in a 5-day FTET protocol. In Experiment 1, 817 dairy heifers were assigned to one of three groups: PGF + estrus detection, 5-d FTET protocol with new (1.38 g P4) or 2nd use CIDR (previously used once for 5 d). Fresh in vitro produced embryos were transferred 7 ± 1 day after estrus (PGF + estrus) or GnRH (5-day FTET). Utilization rate (transferred/treated) was greater (P < 0.001) in heifers submitted to FTET compared to ET after estrus, however pregnancies per ET (P/ET) were not different (P > 0.10). As a result, pregnancy per treated (P/treated) recipient was greater (P < 0.05) in heifers in the 5-day FTET protocol. In Experiment 2, 40 dairy heifers without a corpus luteum (CL) were randomly allocated into one of four groups using new, 2nd use, 3rd use (previously used twice for 5 d/each), or 4th use (previously used thrice for 5 d/each) CIDRs. Circulating P4 was reduced (P < 0.01) with each reutilization. In Experiment 3, ovarian follicular dynamics were evaluated in 238 dairy heifers submitted to a 5-day protocol with either new, 2nd use, 3rd use or 4th use CIDRs at random stages of the estrous cycle. Prostaglandin F2α (PGF) was administered at CIDR removal and again 24 h later. Ovulation was induced by GnRH treatment 72 h after CIDR removal. Preovulatory follicle diameter increased (P < 0.001) progressively with increasing CIDR reutilization. Ovulation rate did not differ between treatments, however, interval from CIDR removal to ovulation decreased (P < 0.001) in heifers receiving 3rd and 4th use CIDRs compared to new or 2nd use. Finally, in Experiments 4 and 5, 1203 heifers submitted to a 5-day FTET protocol were randomly assigned to receive either a new CIDR, a 3rd use CIDR (Experiment 4) or a 4th use CIDR (Experiment 5). Despite the increase in CL volume on D5 in heifers treated with 3rd use (P = 0.03) or 4th use CIDRs (P < 0.01), there were no differences (P > 0.05) in utilization rate, P/ET, or P/treated. Thus, use of a 5-day FTET synchronization protocol improves reproductive efficiency by increasing recipient utilization, and reutilization of CIDRs up to four times in recipient dairy heifers does not compromise reproductive performance.

Hepatic mRNA expression of enzymes associated with progesterone metabolism and its impact on ovarian and endocrine responses in Nelore (Bos indicus) and Holstein (Bos taurus) heifers with differing feed intakes.

The aim of this study was to evaluate circulating progesterone concentration (P4), LH pulsatility and ovarian follicular dynamics in Nelore (B. indicus) and Holstein (B. taurus) heifers under high (HDMI) and low (LDMI) dry matter/energy intakes. In addition, the effects of dry matter/energy intake and breed on hepatic expression of six genes associated with P4 metabolism (AKR1C4, AKR1D1, CYP3A4, CYP2C19, SRD5A1, and SRD5A3) was evaluated. Heifers received an intravaginal P4 device (1 g), 2 mg of estradiol benzoate (EB) i.m. and 500 µg of PGF2α at the begging of the synchronization protocol (D0). Eight days later, the P4 device was removed and all heifers received 1 mg of EB 24h later. Regardless of dry matter/energy intake, the number of recruited follicles was greater in Nelore than in Holstein heifers. In contrast, the maximum diameter of the dominant follicle was greater in Holstein than in Nelore heifers. Circulating P4 concentrations were greater in Nelore than in Holstein from D2 to D9, and in heifers receiving LDMI than those receiving HDMI from D1 to D8 of hormonal protocol. In addition, Holstein heifers had greater LH pulsatility and area under the curve of LH peaks compared to Nelore heifers. However, no effects were observed for LH values between feed intake levels. Interestingly, Holstein heifers had higher expression of SRD5A1, AKR1C4, AKR1D1 than Nelore heifers; whereas, for Nelore heifers, only the expression of CYP3A4 was higher compared to Holstein heifers. In conclusion, there are important differences in the follicular dynamics, circulating P4 and LH pulsatility concentrations that need to be considered during synchronization protocols for Nelore and Holstein breeds. More importantly, these differences appear to be at least partially modulated by the level of feed intake and the contrasting enzyme system in the liver involved with P4 metabolism between these cattle breeds.

Effect of induction of ovulation with estradiol benzoate at P4 device removal on ovulation rate and fertility in Bos indicus cows submitted to a TAI protocol.

This study aimed to minimize the number of times cattle need to be confined during protocols for TAI in beef cows treated for induction of ovulation with EB at the time of P4 device removal (P4r). In Experiment 1, cows were treated with P4 plus EB (Day 0; AM) and were allocated to one of three groups at P4r: EB8.5, EB at P4r on Day 8.5 (PM; three confinements); EB9, EB 24 h after P4r on Day 8 (AM; four confinements) and EC8, EC at P4r on Day 8 (AM; positive control; three confinements). At P4r, cows were treated with PGF2a plus eCG. Ultrasonography was performed from D8 to D12. The interval from P4r to ovulation was less in the EB8.5 compared to EB9 and EC8 group. There was no difference in the ovulation rate between groups. The variability of ovulation was greater in the EB8.5 and EC8 compared to EB9 group. In Experiment 2, cows of EC8 and EB9 groups were submitted to TAI 48 to 52 h (AM) or 54 to 58 h (PM) after P4r (D10). Cows of the EB8.5 group were submitted to TAI 38 to 42 h (AM) or 44 to 48 h (PM) after P4r (D10). There was no interaction between treatments and timing of AI and no treatment effect and timing of AI on P/AI. In conclusion, the delay compared to what typically occurs by 10 h of P4r concomitant with EB administration (Day 8.5) reduced the frequency of animal confinement for the TAI protocol without affecting the reproductive efficiency and the flexibility to perform the TAI in suckled beef cows.

Profile of LH release in response to intramuscular treatment with kisspeptin in Bos indicus and Bos taurus prepubertal heifers.

The main objective of this study was to evaluate the effect of intramuscular (I.M.) administration of different doses of kisspeptin (Kp) on the pattern of luteinizing hormone (LH) release in Bos taurus and Bos indicus prepubertal heifers. Holstein heifers weighing 215.1 ±â€¯38.6 kg (n = 24; aged 5-8 mo) and Gyr heifers weighing 215.4 ±â€¯31.4 kg (n = 24; aged 6-10 mo) were enrolled in this study. The animals were confirmed as non-cyclical by absence of a corpus luteum (CL) as determined by ultrasound scanning of the ovaries and low circulating P4 concentrations (<1.0 ng/mL) evaluated at a 10-day interval (on day -10 and day 0). For each genetic group, heifers were randomly assigned to one of four treatments: Kp at 2.5 µg/kg body weight (Kp2.5), Kp at 5.0 µg/kg (Kp5), Kp at 10 µg/kg (Kp10), or gonadotropin-releasing hormone (GnRH) agonist (0.01 mg of buserelin acetate per heifer), all administered by I.M. injection. All animals responded to the treatments with an LH surge (P < 0.01). There was an effect of breed (P < 0.01) on induced LH release, with Holstein heifers having a greater area under the curve for LH (AUC; P < 0.01) and greater LH peak amplitude (P < 0.01) than Gyr heifers. Nevertheless, greater AUC for LH occurred in Kp10 heifers for both breeds. There was no effect of breed on LH AUC or LH peak amplitude after GnRH agonist treatment. For both breeds, heifers treated with Kp displayed an earlier (P < 0.01) LH peak with a reduced amplitude of the LH peak (P < 0.01) and reduced LH AUC (P < 0.01) compared to heifers treated with GnRH agonist. Thus, both zebu and taurus immature heifers treated with I.M. Kp injection responded with a rapid and dose-dependent LH surge, although even large doses of the native Kp-10 did not mimic the magnitude or duration of the LH surge produced by the GnRH agonist. The early onset of the LH surge after Kp treatment and the short duration suggest that the effects of Kp were likely due to pituitary rather than hypothalamic action. Finally, there seems to be a greater responsiveness to Kp in Bos taurus than in Bos indicus prepubertal heifers, although LH release after GnRH agonist treatment was similar for the two breeds.

Presynchronization by induction of a largest follicle using a progesterone device in GnRH-based-ovulation synchronization protocol in crossbred dairy cows.

The objective was to compare the fertility of dairy cows using a presynchronization protocol by induction of a largest follicle using a progesterone intravaginal device prior to an Ovsynch protocol (P4synch) with the Double-Ovsynch in lactating dairy cows. Lactating Bos indicus x Bos taurus crossbred cows (n = 440) were randomly allocated to one of two treatments: (I) Double-Ov (n = 228), GnRH (D-17), PGF2α 7 days later (D-10) and GnRH 3 days later (D-7) followed by an Ovsynch protocol 7 days later (GnRH on D0, PGF on D7, GnRH on D9); (II) P4synch (n = 212), insertion of a sustained release progesterone intravaginal device (D-10), 10 days later (D0) an Ovsynch protocol was initiated (GnRH on D0, PGF on D7, GnRH on D9) with progesterone device withdrawal on Day 7. All cows were artificially inseminated (TAI) 16 h after the second GnRH of the Ovsynch protocol and pregnancy diagnosis was performed by transrectal ultrasonography 30 and 60 days after TAI. A subset of cows (n = 52 for Double-Ov and n = 50 for P4synch) ultrasonography was performed on days 0, 7, 9 and 24 of the experimental period. There were no differences among treatments on presynchronization rate [presence of a follicle>12 mm on D0, Double-Ov 94.2% (49/52) and P4synch 92.0% (46/50); P = 0.66], follicular diameter on the 1st GnRH (Double-Ov 17.2 ±â€¯0.7 mm e P4synch 18.6 ±â€¯0.9 mm; P = 0.28), ovulation rate to the 1st GnRH [Double-Ov 86.3% (44/51) and P4synch 81.2% (39/48); P = 0.50], synchronization rate [presence of a follicle>12 mm on D9; Double-Ov 84.6% (44/52) and P4synch 86.0% (43/50); P = 0.84], follicular diameter on the 2nd GnRH (Double-Ov 17.5 ±â€¯0.6 mm and P4synch 18.0 ±â€¯0.5 mm; P = 0.48), ovulation rate to the 2nd GnRH [Double-Ov 90.9% (40/44) and P4synch 86.0% (37/43); P = 0.48] and CL diameter on D24 (Double-Ov 27.9 ±â€¯0.7 mm and P4synch 29.4 ±â€¯0.9 mm; P = 0.19). Corpus luteum presence on D0 was different (P = 0.03) among treatments [Double-Ov 57.7% (30/52) and P4synch 36.0% (18/50)]. There was no difference (P = 0.85) among the pregnancy per AI on day 30 [Double-Ov 39.0% (89/228) and P4synch 40.1% (85/212)], on day 60 [Double-Ov 34.8% (79/227) and P4synch 38.7% (82/212); P = 0.41] and pregnancy loss [Double-Ov 10.2% (9/88) and P4synch 3.5% (3/85); P = 0.08]. The presynchronization by induction of a largest follicle using a sustained release progesterone device prior to Ovsynch yielded similar results compared with the Double Ovsynch protocol on follicular development patterns and on the fertility of lactating dairy cows.

Hormonal strategy to reduce suckled beef cow handling for timed artificial insemination with sex-sorted semen.

Two experiments were conducted to assess a hormonal strategy developed to reduce animal handling for timed artificial insemination (TAI) with sex-sorted semen. Four-hundred ninety-one (491) suckled beef cows received a progesterone (P4) intravaginal device and 2 mg intramuscular (im) injection of estradiol benzoate (EB) on a randomly chosen day of the estrus cycle (Day 0) in Experiment 1. Cows were treated with 500 µg of sodic cloprostenol (PGF2α) and with 300 IU of eCG at P4 device removal (Day 8); these cows were also randomly assigned to receive 1 mg of estradiol cypionate (EC) administered at P4 device removal (treatment EC-0h) or 1 mg of EB 24 h after P4 device removal (treatment EB-24h). Both treatments were timed inseminated (TAI) with sex-sorted semen 60 h after P4 device removal. Cows treated with EC-0h presented higher pregnancy rate per AI (P/AI) [45.0% (113/251)] than the ones treated with EB-24h [35.4% (85/240); P = 0.03)]. A subset of cows (n = 26) were subjected to ultrasound examination every 12 h after P4 device removal for 96 h in the row in order to determine the time of ovulation. Similar interval between device removal and ovulation was recorded for EB-24h = 70.0 ±â€¯2.9 h vs. EC-0h = 66.0 ±â€¯2.8 h (P = 0.52). Five-hundred ninety-one (591) cows were subjected to the same synchronization protocols and treatments (EC-0h or EB-24h). In addition, they were randomly assigned to a 2 × 2 factorial arrangement aiming at determining the effects of treatment with estradiol (EC-0h vs. EB-24h) and of semen type (Sex-sorted vs. Non-sex-sorted semen). All animals were timed inseminated 60 h after P4 device removal. There was no interaction (P = 0.07) between the ovulation inducer and semen type. The EC protocol led to greater P/AI than EB (P = 0.03). Greater (P = 0.01) P/AI was achieved through treatments with non-sex-sorted semen rather than with sex-sorted semen [sex-sorted (EB-24h = 49.0%; EC-0h = 51.0%) vs. non-sex-sorted semen (EB-24h = 52.4%; EC-0h = 68.2%)]. Therefore, EC administered at P4 device removal resulted in greater P/AI. Furthermore, the EC-0h protocol allowed reducing suckled beef cow handing for timed artificial insemination with sex-sorted semen.

Synchronization treatments previous to natural breeding anticipate and improve the pregnancy rate of postpartum primiparous beef cows.

Timed AI has become a potential tool to bypass postpartum acyclicity, yet only a small percentage of the world bovine herd is inseminated. Most females are still subjected to bull mating; therefore, the frequent occurrence of postpartum anestrus may compromise their reproductive efficiency. Thus, the aim of this study was to develop an approach that allows the early conception of postpartum primiparous beef cows that are exposed to natural breeding (NB). For this purpose, 350 primiparous Nelore cows 35-65 d postpartum were allocated into three groups: Control (no hormonal treatment), TNB (hormonal protocol for timed-NB without equine chorionic gonadotropin; eCG) and TNB + eCG (hormonal protocol for TNB with eCG) groups. The protocol for TNB consisted of the insertion of a 1-g progesterone device and the intramuscular (IM) administration of 2 mg estradiol benzoate on D-9 (nine days before bull exposure), followed by device removal and the administration of 1 mg estradiol cypionate IM on D0. An additional 300 IU of eCG was given only to TNB + eCG cows. All cows were exposed to bull mating from D0 to D105. Pregnancy was checked by ultrasonography every 50 d, gestational age was estimated, and the number of new gestations every 21-d cycle (P21, P42, P63, P84 and P105) was predicted. Control cows had lower pregnancy at P21 (5.7%c, 7/123) than TNB (30.4%b, 35/115) and TNB + eCG (51.8%a, 58/112; P = 0.001) cows. Pregnancy rate increased across P42, P63 and P84 (P = 0.001) but remained higher for TNB + eCG cows. Regarding time to conception, the TNB + eCG cows achieved a greater than 50% pregnancy rate within the first days after bull exposure, while the TNB and Control cows took more than 40 d and 90 d to achieve 50% pregnancy rates, respectively. At the end of the breeding season (BS), TNB + eCG cows had a 21% higher pregnancy rate than the Control cows and a 16% higher rate than the TNB cows. The probability of conceiving increased 1.5-fold for cows treated with TNB (P = 0.0079) and 2.2-fold for cows additionally treated with eCG (P < 0.0001). The average interval between the onset of the BS and conception was reduced (P < 0.0001) for the TNB + eCG cows (26.5 ± 3.8c) compared to the TNB (35.7 ±â€¯4.1b) and Control (64.7 ±â€¯3.9a days) cows. Thus, the use of TNB, especially when associated with eCG, efficiently improved the early conception of postpartum primiparous beef cows after exposure to NB. The increased number of cows conceiving early in the BS is crucial to improve reproductive efficiency by reducing the interval between parturitions and improving the number of pregnant cows at the end of BS, thus resulting in greater farm income.

Review: Using artificial insemination v. natural service in beef herds.

The aim of this review is to compare the performance of different reproductive programs using natural service, estrus synchronization treatment before natural service (timed natural breeding (TNB)), artificial insemination (AI) following estrus detection and timed artificial insemination (TAI) in beef herds. It is well known that after parturition the beef cow undergoes a period of anestrous, when they do not exhibit estrus, eliminating the opportunity to become pregnant in the early postpartum by natural mating or by AI after detection of estrus. Hormonal stimulation is already a consistent and well-proven strategy used to overcome postpartum anestrus in beef herds. Basically, hormones that normally are produced during the estrous cycle of the cow can be administered in physiological doses to induce cyclicity and to precisely synchronize follicular growth, estrus and ovulation. Furthermore, two options of mating may be used after hormonal stimulation: natural service (i.e. utilization of bull service after synchronization, referred to as TNB) and TAI. These strategies improve the reproductive efficiency of the herds compared with natural service without estrus induction or synchronization. After the first synchronized service, the most common strategy adopted to get non-pregnant cows pregnant soon is the introduction of clean-up bulls until the end of the breeding season. However, methods to resynchronize non-pregnant cows after the first service are already well established and offer a potential tool to reduce the time for subsequent inseminations. Thus, the use of these technologies enable to eliminate the use of bulls by using resynchronization programs (i.e. two, three or four sequential TAI procedures). The dissemination of efficient reproductive procedures, such as TNB, TAI and Resynch programs, either isolated or in combination, enables the production of a greater quantity (obtaining increased pregnancy rates early in the breeding season) and quality (maximization of the use of AI with superior genetic sires) of beef calves. These technologies can contribute to improve the production efficiency, and consequently, improve livestock profitability.

The effect of circulating progesterone on magnitude of the GnRH-induced LH surge: Are there any differences between Bos indicus and Bos taurus heifers?

This study evaluated the effects of differing circulating progesterone (P4) levels on the luteinizing hormone (LH) surge profile following treatment with gonadotropin releasing hormone (GnRH) in B. indicus (Nelore, n = 13) and B. taurus (Holstein, n = 16) heifers. All heifers were synchronized with a hormonal protocol to induce either a Low or High circulating P4 environment at the time of GnRH treatment. Heifers were randomly assigned to a 2 × 2 factorial design composed by two genetic groups (B. indicus and B. taurus) and two levels of circulating P4 concentrations (Low or High). Blood samples were collected every 30 min from -30 to 210 min and at 270 min after GnRH treatment. As expected, mean P4 concentration was greater for cows in the High than in the Low P4 group (P = 0.0008) and in Bos indicus than in Bos taurus heifers (P = 0.06). Despite genetic group, the area under the curve of LH release was greater in the Low-P4 than in High-P4 concentration group (P < 0.0001). Interestingly, it appears that High P4 concentrations had a more pronounced effect on LH peak in B. indicus than in B. taurus heifers, as indicated by the interaction (P = 0.01) between genetic group and P4 levels. In conclusion, circulating P4 concentration have a great impact on the GnRH-induced LH surge of both B. indicus and B. taurus heifers, but it does not explain the much lower LH peak in B. indicus with low circulating P4. Thus, more studies are essential to uncover some of the underlying physiological factors other than circulating P4 that are limiting LH release following a GnRH treatment in B. indicus cattle.

Beef donor cows with high number of retrieved COC produce more in vitro embryos compared with cows with low number of COC after repeated ovum pick-up sessions.

The aim of the present study was to evaluate whether repeated ovum pick-up (OPU) procedures may affect the efficiency of OPU programs for in vitro embryo production (IVEP) in Bos indicus (Nelore) donors. In addition, the repeatability (r) efficiency of IVEP was also assessed. Data available were from 432 OPU-IVEP sessions that were performed at random stages of the estrous cycle in 36 cycling, nonlactating Nelore donors. Semen from three Nelore bulls was used for the IVF. Donors were submitted to 12 consecutive OPU procedures, with an interval of approximately 30 days between sessions. Data were analyzed as repeated measures using the GLIMMIX procedure of SAS 9.3. Cows yielding ≥15 COCs were defined as "high" COCs and cows with less than 15 COCs were defined as "low" COCs donors. The number of COCs retrieved decreased over time in donors classified with high COCs and remained fairly steady in cows with low COCs at the beginning of the program (Psession*COC retrieved groups = 0.02). Moreover, the number of COCs retrieved (P < 0.0001), and number of blastocysts produced per OPU (P = 0.001) was greater for the high COCs donors compared with the low COCs category, and these results were consistent across OPU sessions. Interestingly, there was no effect of COCs category on the rate of blastocyst development (P = 0.83). In addition, number of blastocysts produced were not affected by repeated OPU (P = 0.37) and interactions between time and COC category (P = 0.72). Similarly, blastocyst rate was not affected by repeated OPU (P = 0.21) and interactions between time and COC category (P = 0.58). Despite of COC category of the donor cows, repeatability was high for the number of COCs retrieved (r = 0.81), number of blastocysts produced per OPU (r = 0.79), and blastocyst rate (0.69). In conclusion, overall numbers of COCs decreased over time in donors classified as having high COCs. However, cumulative amounts of produced blastocysts were greater in donors with high COCs. More importantly, high repeatability was observed in terms of IVEP efficiency. Therefore, IVEP programs can be significantly improved with more aggressive selection toward donors with greater numbers of COCs.

Inducing ovulation with oestradiol cypionate allows flexibility in the timing of insemination and removes the need for gonadotrophin-releasing hormone in timed AI protocols for dairy cows.

The effects of addition of gonadotrophin-releasing hormone (GnRH) to a progesterone plus oestradiol-based protocol and timing of insemination in Holstein cows treated for timed AI (TAI) were evaluated. Cows (n=481) received a progesterone device and 2mg oestradiol benzoate. After 8 days, the device was removed and 25mg dinoprost was administered. Cows were allocated to one of three (Study 1; n=57) or four (Study 2; n=424) groups, accordingly to ovulation inducer alone (Study 1; oestradiol cypionate (EC), GnRH or both) or ovulation inducer (EC alone or combined with GnRH) and timing of insemination (48 or 54h after device removal; Study 2). In Study 1, the diameter of the ovulatory follicle was greater for GnRH than EC. Oestrus and ovulation rates were similar regardless of ovulatory stimuli. However, time to ovulation was delayed when GnRH only was used. In Study 2, cows treated with GnRH or not had similar pregnancy per AI (P/AI) 30 days (41.5% vs 37.3%; P=0.28) and 60 days (35.9% vs 33.0%; P=0.61) after TAI. TAI 48 and 54h after device removal resulted similar P/AI at 30 days (40.3% vs 38.5%; P=0.63) and 60 days (33.8% vs 35.1%; P=0.72). Thus, adding GnRH at TAI does not improve pregnancy rates in dairy cows receiving EC. The flexibility of time to insemination enables TAI of a large number of cows using the same protocol and splitting the time of AI.

Ovulation synchronization with estradiol benzoate or GnRH in a timed artificial insemination protocol in buffalo cows and heifers during the nonbreeding season.

The aim of this study was to compare estradiol benzoate (EB) and GnRH for the induction of ovulation in a TAI protocol in buffalo during the nonbreeding season. In experiment 1, 141 buffaloes (56 cows and 85 heifers) received an intravaginal P4 device (1.0 g) plus EB (2.0-mg, intramuscular [im]) at random stage of the estrous cycle (Day 0). On Day 9, the P4 device was removed, and buffaloes were given PGF2α (0.53-mg im sodium cloprostenol) plus eCG (400-IU im). Buffaloes were then randomly allocated to one of three groups and treated as follows: EB24 (n = 47), EB (1.0 mg im) 24 hours after P4 device removal; EB36 (n = 50), EB 36 hours after P4 device removal; GnRH48 (n = 44), GnRH (10 µg im buserelin acetate) 48 hours after P4 device removal. Ultrasound examinations were performed on Day 0 to ascertain ovarian follicular status, Day 9 to measure follicular diameter, and from Day 11 to Day 14 (every 12 hours for 60 hours) to establish the time of ovulation. There were no significant differences between EB24, EB36, and GnRH48 for diameter of the ovulatory follicle (13.1 ± 0.3, 13.7 ± 0.3, and 13.7 ± 0.3 mm; P = 0.26) and ovulation rate (78.7%, 82.0%, and 84.1%; P = 0.93). When compared with heifers, cows had a greater diameter of the dominant follicle on Day 9 (10.3 ± 0.3 and 8.6 ± 0.2 mm; P = 0.0001), diameter of the ovulatory follicle (14.1 ± 0.3 and 13.1 ± 0.2 mm; P = 0.01), ovulation rate (91.1% and 75.3%; P = 0.02), and interval from P4 device removal to ovulation (76.3 ± 1.3 and 72.5 ± 1.4 hours; P = 0.05). In experiment 2, 511 buffaloes (354 cows and 157 heifers) were assigned to the same treatments described in experiment 1 (EB24, n = 168; EB36, n = 172; and GnRH48, n = 171), and all animals were submitted to timed artificial insemination (TAI) 64 hours after P4 device removal. Pregnancy diagnosis was undertaken 30 days after TAI. There were no significant differences between EB24, EB36, and GnRH48 for pregnancy rate (45.2%, 43.0%, and 49.7%; P = 0.46), and the pregnancy rate did not differ (P = 0.31) for cows (47.5%) and heifers (42.7%). The findings from the two experiments indicated that EB (24 or 36 hours) and GnRH (48 hours) induce comparable follicular responses, ovulation, and pregnancy rates in buffalo cows and heifers. Although there were some differences in the follicular responses between cows and heifers, the pregnancy rate to TAI was nonetheless similar.

Effects of eCG are more pronounced in primiparous than multiparous Bos indicus cows submitted to a timed artificial insemination protocol.

The effects of eCG on follicular growth, ovulation, and pregnancy per artificial insemination (P/AI) in multiparous and primiparous Bos indicus beef cows submitted to timed artificial insemination (TAI) were evaluated in three experiments. In experiments 1 (follicular responses; n = 64), 2 (follicular growth and ovulation rate; n = 662), and 3 (P/AI; n = 2092), cows submitted to TAI were assigned to receive one of two treatments on Day 8 of the synchronization protocol: control (no additional treatment) or eCG (300-IU of eCG intramuscularly). In experiment 1, largest follicle (LF) diameter on Day 8 (P = 0.56) and the interval from progesterone (P4)-device removal to ovulation (P = 0.79) did not differ between treatments. However, the maximum diameter of the LF (P = 0.05) and ovulation rate (P = 0.03) were greater in cows that received eCG. In experiment 2, the diameter of the LF on Day 10, follicular growth, and ovulation rate were greater in eCG-treated cows (P < 0.01). However, CL diameter was similar between treatments (P = 0.11). In experiment 3, there was a treatment-by-parity interaction (P = 0.003) on P/AI, such that treatment with eCG was more effective in primiparous cows. In conclusion, eCG treatment resulted in increased final follicular growth, ovulation rate, and fertility in B indicus cows submitted to TAI protocols, especially in primiparous cows.