Bovine oviductal fluid (bOF) collected in the follicular or luteal phase of the estrous cycle exerts similar effects on ram sperm kinematics and acrosome reactivity in vitro.

This study examined the effects of bovine oviductal fluid (bOF) obtained during the follicular or luteal phase of the estrous cycle on ram sperm kinematics, capacitation status and plasma membrane (PM) integrity at various time points during the 24-h incubation period. Fresh ram spermatozoa were selected using the swim-up technique and then incubated separately with either follicular phase (FbOF) or luteal phase (LbOF) bovine oviductal fluid added to Fert-TALP medium (positive control - POSControl) or in Fert-TALP medium without capacitating agents (negative control - NEGControl) at 38 °C under 5% CO2. Incubation with FbOF or LbOF for 2 h and 4 h promoted an increase (P <  0.05) in most of the sperm motility parameters as compared with the NEGControl group, and bOF-induced changes in sperm kinematics were similar (P >  0.05) to those seen in the POSControl group. After 6 h of incubation, the stimulatory effect of FbOF or LbOF on ram sperm kinematics was no longer observed (P >  0.05). Sperm PM integrity was not affected (P >  0.05) by incubation in bOF-supplemented media or in absence of capacitating factors (NEGControl). Although neither FbOF nor LbOF had any effect on sperm capacitation rates, the proportion of acrosome-reacted spermatozoa was greater (P < 0.05) for bOF-containing media compared with the NEGControl group during the long incubation periods (18 h and 24 h). In conclusion, bOF from either follicular or luteal phase of the estrous cycle enhances ram sperm motility for up to 4 h and the rate of acrosome reaction after long (18-24 h) incubation periods without affecting sperm viability.

Anti-Müllerian hormone and antral follicle count are more effective for selecting ewes with good potential for in vivo embryo production than the presence of FecGE mutation or eCG pre-selection tests.

This study aims to compare four different methods for selecting high responding sheep donors for in vivo embryo production. These methods include a pre-selection eCG test (eCG), antral follicle count (AFC), plasma anti-Müllerian hormone measurement (AMH) and genotyping for the presence of the FecGE mutation (a polymorphism in the GDF9 gene associated with increased ovulation rate). Santa Ines ewe lambs (n = 25) underwent superovulation (SOV) with 800 IU equine chorionic gonadotropin (eCG) and the corpus luteum (CL) count was recorded by laparoscopy after eight days. At the D0eCG, blood samples for AMH and genotyping analysis were collected. Twenty-one days after the end of the eCG test, the same animals underwent SOV with 200 mg of FSH, administered in six decreasing doses, and then naturally mated. Immediately before the beginning of the FSH protocol (D0FSH), and at the moment of the first FSH dose (D9FSH), the AFC was assessed. Plasma AMH was again determined at the D9FSH. After each screening process, animals were classified as having a high (HR), or low (LR), potential of response (using specific thresholds for each method). Then, the ewes' response to SOV and embryo yield for each screening method, classified as HR or LR, were compared. Animals classified as HR by AFC (HRAFC) and by AMH concentration (HRAMH) at the D9FSH, produced more viable embryos than those classified as LRAFC and LRAMH (HRAFC 6.2 ±â€¯3.2 vs LRAFC 2.8 ±â€¯3.0 and HRAMH 6.6 ±â€¯3.6 vs LRAMH 3.0 ±â€¯2.9). Pre-selection tests with eCG and different FecGE genotypes, either heterozygous (+/E) or wild type (+/+), were unable to discriminate HR or LR animals. A tendency (P = 0.06) to have lower plasma AMH was observed in heterozygous FecGE (+/E) ewes. In conclusion, both AFC and plasma AMH can be used to select donor ewes with a higher potential of response for in vivo embryo production.

Early pregnancy diagnosis in ewes by subjective assessment of luteal vascularisation using colour Doppler ultrasonography.

This study was designed to evaluate early pregnancy diagnosis using colour Doppler ultrasonography (US) for luteal vascularisation assessment. In Study 1, 28 ewes were artificially inseminated (Day 0), and luteal vascularisation was assessed from Day 12 to Day 19 by two evaluators using colour Doppler US, categorising the corpus luteum (CL) on a subjective scale ranging from 1 to 4. Females bearing a CL with score 2 or greater were presumably considered pregnant. Pregnancy was confirmed on Day 30 by B-Mode US. In Study 2, a predictive pregnancy diagnosis was performed on Day 17 in 197 ewes based on the criteria described in Study 1. Pregnancy was confirmed by B-mode US on Day 45. Agreement between evaluators was verified by an intraclass correlation coefficient (ICC) and Kappa index (κ). Performance of colour Doppler US for early pregnancy diagnosis was evaluated calculating sensitivity (Sens), specificity (Spec), negative predictive values (NPV), positive predictive values (PPV) and accuracy (Ac). In Study 1, luteal vascularisation assessment was unable to predict non-pregnant animals between 12 and 14 days after insemination, as all animals still had vascularised CL, and thus were considered pregnant. The colour Doppler US performance improved progressively until Day 17, when it reached maximum values (Sens = 100%, Spec = 76%, PPV = 73%, NPV = 100% and Ac = 86%). The subjective scale for luteal irrigation assessment showed medium to good agreement among evaluators on Day 12 and Day 13 (ICC = 0.66 and 0.68, respectively), and excellent agreement from Day 14 to Day 19 (ICC = 0.90, 0.80, 0.80, 0.84, 0.95 and 0.93, respectively). Agreement was almost perfect for score 1 CLs (κ = 0.87), and moderate for scores 2, 3 and 4 CLs (κ = 0.54, 0.48 and 0.41, respectively). In Study 2, performance of colour Doppler US as a tool to predict pregnancy status in ewes on Day 17 post-insemination was as follows: Sens = 93.5%, Spec = 80.8%, PPV = 85.6%, NPV = 91.1% and Ac = 87.8%. Subjective luteal vascularisation assessment using colour Doppler US to distinguish between pregnant and non-pregnant animals was considered a reliable tool which was highly efficient beginning 17 days after breeding.

Use of two doses of cloprostenol in different intervals for estrus synchronization in hair sheep under tropical conditions.

This study evaluated the effect of two doses of prostaglandin at different intervals on reproductive parameters of crossbred ewes. In Experiment 1, 30 ewes received two doses of 120 µg cloprostenol at 7 (G 7 days), 9 (G 9 days), or 11.5 (G 11.5 days) days apart. Ultrasound assessments were performed from the first and second cloprostenol administration for 5 days or ovulation detection. Estrus signs were checked by a teaser male. Plasma progesterone concentration was measured before each cloprostenol dose. In Experiment 2, 95 ewes were allocated into the same treatments and after the second dose, ewes in estrus were mated. At 30 days after breeding, pregnancy diagnosis was conducted and prolificacy was evaluated at lambing. In Experiment 1, at the first cloprostenol administration, 50% of ewes had an active CL and all showed estrus. At the second administration, 66.7% of ewes had an active CL and one did not present estrus. There was no difference (P > 0.05) after the second dose for as follows: overall estrous response (90%), interval from cloprostenol administration to estrous onset (42.0 ± 4.9 h), estrus duration (31.5 ± 2.1 h), ovulation rate (100.0%), and number of ovulations (1.5 ± 0.3). In Experiment 2, both pregnancy and prolificacy rates were similar (P > 0.05) for G 7 days (73.3; 145%), G 9 days (75.9; 125%), or G 11.5 days (75.9; 145%), leading to an overall pregnancy rate of 75.0% (66/88) and prolificacy rate of 137%. Therefore, the three treatments proposed were able to promote high pregnancy and prolificacy rates in crossbred ewes.