Transcervical uterine flushing and embryo transfer in sheep: Morphophysiological basis for approaches currently used, major challenges, potential improvements, and new directions (alas, including some old ideas).

At present, the success of non-surgical embryo recovery (NSER) and transfer (NSET) hinges upon the cervical passage of catheters, but penetration of the uterine cervix in ewes is problematic due to its anatomical structure (i.e., long and narrow cervical lumen with misaligned folds and rings). It is a major obstacle limiting the widespread application of NSER and NSET in sheep. While initial attempts to traverse the uterine cervix focused on adapting or re-designing insemination catheters, more recent studies demonstrated that cervical relaxation protocols were instrumental for transcervical penetration in the ewe. An application of such protocols more than tripled cervical penetration rates (currently at 90-95 %) in sheep of different breeds (e.g., Dorper, Lacaune, Santa Inês, crossbred, and indigenous Brazilian breeds) and ages/parity. There is now sufficient evidence to suggest that even repeatedly performed cervical passages do not adversely affect overall health and reproductive function of ewes. Despite these improvements, appropriate selection of donors and recipients remains one of the most important requirements for maintaining high success rates of NSER and NSET, respectively. Non-surgical ovine embryo recovery has gradually become a commercially viable method as even though the procedure still cannot be performed by untrained individuals, it is inexpensive, yields satisfactory results, and complies with current public expectations of animal welfare standards. This article reviews critical morphophysiological aspects of transcervical embryo flushing and transfer, and the prospect of both techniques to replace surgical methods for multiple ovulation and embryo transfer (MOET) programs in sheep. We have also discussed some potential pharmacological and technical developments in the field of non-invasive embryo recovery and deposition.

Oocyte Competence of Prepubertal Sheep and Goat Oocytes: An Assessment of Large-Scale Chromatin Configuration and Epidermal Growth Factor Receptor Expression in Oocytes and Cumulus Cells.

The oocyte competence of prepubertal females is lower compared to that of adults, mainly because they originate from small follicles. In adult females, the germinal vesicle (GV) and epidermal growth factor receptor (EGFR) have been associated with oocyte competence. This study aimed to analyze GV chromatin configuration and EGFR expression in prepubertal goat and sheep oocytes obtained from small (<3 mm) and large (≥3 mm) follicles and compare them with those from adults. GV chromatin was classified from diffuse to condensed as GV1, GVn, and GVc for goats and NSN, SN, and SNE for sheep. EGFR was quantified in cumulus cells (CCs) by Western blotting and in oocytes by immunofluorescence. Oocytes from prepubertal large follicles and adults exhibited highly condensed chromatin in goats (71% and 69% in GVc, respectively) and sheep (59% and 75% in SNE, respectively). In both species, EGFR expression in CCs and oocytes was higher in prepubertal large follicles than in small ones. In adult females, EGFR expression in oocytes was higher than in prepubertal large follicles. In conclusion, GV configuration and EGFR expression in CCs and oocytes were higher in the large than small follicles of prepubertal females.

The presence of estradiol benzoate in the cervical relaxation treatment for non-surgical embryo collection does not impair embryonic morphological quality, cryosurvival, and gene expression profile.

Non-surgical embryo recovery (NSER) is usually preceded by a cervical relaxation in ovine donors, based on estradiol benzoate (EB), prostaglandin (PGF), and oxytocin (OT). However, it is hypothesized that, due to poorly understood mechanisms, EB can result in embryotoxic actions. To evaluate this, 20 min before NSER superovulated sheep were induced to cervical relaxation with 0.0 (G0.0), 0.5 (G0.5), or 1.0 mg (G1.0) of EB associated with 37.5 µg of PGF 16 h before NSER and 50 IU of OT. In doing so, the efficiency and duration of the NSER procedure showed no compromise (P > 0.05). Additionally, the presence of EB did not affect (P > 0.05) the embryo's morphological quality, the development dynamics, or the abundance of transcripts associated with embryonic quality (OCT4 and NANOG), cellular stress (HSP90 and PRDX1), and apoptosis (BCL2 and BAX). A similar result (P > 0.05) was also observed when comparing embryonic cryosurvival at 24 (52.0, 52.0, and 54.0) and 48 h (60.0, 54.0, and 58.0) of in vitro culture (G0.0, G0.5, and G1.0, respectively). Thus, we can conclude that EB use does not compromise embryonic quality and cryoresistance.

Luteal tissue characteristics of Morada Nova ewes with hCG application 7.5 days after the end of estrus synchronization protocol in the breeding season.

Results with the use of hCG after synchronization protocol are still inconsistent, which may vary according to breed, season, day of application and dose of the drug used. The aim of the present study was to evaluate the functionality of luteal tissue and ovarian perfusion after hCG treatment during early luteal phase. Estrus-synchronized ewes were randomly assigned to receive i.m. injection of 300 IU of hCG (G-hCG; n = 40) or 1 mL of saline (G-Control; n = 32) on Day 7.5 after progesterone withdrawal. Ultrasonographic evaluations of the ovaries and ovarian and iliac arteries were performed on Days 7.5, 10.5, 13.5, and 21.5. The accessory corpus luteum (aCL) formation rate was 52.5% for G-hCG. There was interaction (p > 0.05) for treatment (G-hCG and G-Control), days (7.5, 10.5, 13.5 and 21.5) and PD (Pregnant and Non-pregnant) for the variables of biometric characteristics of the corpus luteum B-Mode and Color Doppler on days 7.5, 10.5, 13.5 and 21.5. There was no difference (p > 0.05) for pregnancy rates and mean fetuses per ewe between the treatment groups. It is concluded that the application of hCG 7.5 days after the hormonal protocol in Morada Nova ewes in a breeding season is efficient in inducing aCL formation and increasing luteal tissue biometry. However, there was no effect on pregnancy rate.

Transcervical artificial insemination in the brown brocket deer (Subulo gouazoubira): a promising method for assisted reproduction in deer.

The present study aimed to test the efficiency of transcervical artificial insemination techniques with cervical immobilization (TCAI-CI) or cervical traction (TCAI-CT), associated or not with the use of oxytocin (OT) as a protocol for cervical dilation, in the brown brocket deer (Subulo gouazoubira). The study was carried out in a crossover design using four adult females in two replicates with an interval of 60 days. Estrus was synchronized with oral melengestrol acetate (MGA) associated with estradiol benzoate and sodium cloprostenol. TCAI techniques were performed from 18 to 24 h after estrus onset. All females received either an i.v. application of 50 IU of OT (G-OT, n = 4) or 1 mL of saline solution (G-Control, n = 4) 20 min before the TCAI procedure. The TCAIs were performed using frozen-thawed semen motility 40%, vigor 3, acrosome integrity 87%, membrane integrity of 95% and 13% of total post-thaw defects from the same batch. Behavioral estrus was observed in 100% of the females, in both replicates. It was achieved a 50% (4/8) success of cervical transposition with semen deposition in the uterine. Regarding inseminations, most of them (87.5%) were performed using the TCAI-CT technique, and the overall conception rate was 50%. Cervical transposition times (< 1 min) and TCAI procedures (~ 17 min) were considered satisfactory. Thus, the performance of the TCAI-CI and TCAI-CT techniques was successful, regardless of using OT as a cervical dilation protocol. This procedure is proposed as a method of choice for artificial insemination with greater applicability in different conservation centers, compared to more advanced reproductive biotechniques, and with a favorable impact on the conservation of deer species.

An Exploratory Data Analysis from Ovine and Bovine RNA-Seq Identifies Pathways and Key Genes Related to Cervical Dilatation.

The present study developed a review and exploration of data in public and already validated repositories. The main objective was to identify the pathways involved in ruminants' cervical dilatation, which are conserved between cattle and sheep in the follicular and luteal phases of the reproductive cycle. In cattle, 1961 genes were more differentially expressed in the follicular phase and 1560 in the luteal phase. An amount of 24 genes were considered exclusively expressed from these. A total of 18 genes were in the follicular phase and 6 genes were in the luteal phase. In sheep, 2126 genes were more differentially expressed in the follicular phase and 2469 genes were more differentially expressed in the luteal phase. Hoxb genes were identified in both species and are correlated with the PI3K/Akt pathway. PI3K/Akt was also found in both cattle and sheep, appearing prominently in the follicular and luteal phases of both species. Our analyses have pointed out that the PI3K/Akt pathway and the Hoxb genes appear in prominence in modulating mechanisms that involve estrus alterations in the cervix. PI3K/Akt appears to be an important pathway in the cervical relaxation process.

Climatic seasons and time of the day influence thermoregulation and testicular hemodynamics in Santa Inês rams raised under humid tropical conditions.

This study evaluated the possible association between the diurnal variations of climatic factors during the rainy (RS) or less rainy (LS) seasons on the testicular hemodynamics and thermoregulatory responses of hair sheep rams raised in a humid tropical climate. Santa Inês rams (n = 6) underwent evaluation of general and testicular physiological parameters (heart and respiratory rates, internal and scrotal temperatures, internal-scrotal temperature gradient, scrotal distention, and color Doppler ultrasound evaluation of the spermatic cords and spectral analyses of testicular arteries) over six consecutive weeks per season at three separate times daily (morning = 8:00 a.m., noon = 12:00 p.m., and afternoon = 5:00 p.m.) during the RS and LS. Climatic air temperature and relative humidity data were recorded, and the temperature and humidity index (THI) was calculated. Higher thermal challenge was observed in LS relative to RS (air temperature = 28.0 vs. 30.9 °C; relative humidity = 84.1 vs. 69.9%; THI = 80.0 vs. 82.5; P < 0.05). In both seasons, respiratory rate and internal temperature were normal, demonstrating the animals' adaptability. In RS, however, a higher scrotal temperature was recorded in relation to LS (35.0 vs. 34.7 °C; P < 0.05), with a gradual increase from morning to afternoon. Lower resistivity (0.40 vs. 0.64; P < 0.05) and pulsatility (0.55 vs. 1.14; P < 0.05) indices, and a higher rate of high-velocity blood flow of testicular arteries (71.1 vs. 60.6%; P < 0.05) were observed in RS compared to LS. The lowest correlations between testicular hemodynamic, physiological variables, and environmental parameters (P < 0.05) were observed in the morning. In conclusion, testicular thermoregulation and testicular hemodynamics were influenced by the climatic seasons and time of the day, being more efficient in the LS season and with less interference from environmental factors in the morning.

Intravaginal progesterone device reinsertion during the early luteal phase affects luteal function and embryo yield in superovulated ewes.

This study checked the efficacy of progesterone (P4) device reinsertion during the early luteal phase on luteal function and embryo yield in superovulated crossbred ewes. Twenty multiparous ewes received an intravaginal P4 device for nine days (D0 to D9) followed by six decreasing doses (25, 25, 15, 15, 10, 10%) of 133 mg pFSH i.m. at 12 h intervals, starting 60 h before P4 device removal. Ewes were naturally mated at 12 h intervals while in estrus. On D13, ewes with viable corpora lutea (CL; n = 19) were equally allocated for receiving their P4 device reinsertion (G-P4; n = 10) or not (G-Control; n = 9). On D17, the P4 device was removed, and all females received the cervical relaxation protocol 16 h to 20 min before non-surgical embryo recovery. CL count and their functionality classification were performed on D13 and D17 by transrectal B-mode and color Doppler ultrasonography (US). Plasma P4 concentrations (ng/mL) of G-P4 ewes increased (P < 0.05) over the days, being greater on D17 (9.2 ± 2.8) than on D9 (1.9 ± 0.7) and D13 (1.6 ± 0.4). The overall CL count per ewe tended to be greater (P = 0.09) in G-P4 compared with G-Control. The occurrence of premature regression of corpora lutea did not differ (P > 0.05) between G-P4 (30.0%) and G-Control (44.4%). The number of ova/embryos recovered was greater (P < 0.05) in G-P4 (11.6 ± 2.9) compared with G-Control (3.7 ± 2.0), respectively. Altogether, the reinsertion of the P4 device for four days after superovulation in ewes promotes greater P4 concentrations, resulting in greater ova/embryos recovered.

Luteal Function, Biometrics, and Echotextural Attributes in Santa Inês Ewes Superovulated with Different Total Doses of Porcine Follicle-Stimulating Hormone.

Premature regression of corpora lutea (PRCL) may adversely affect the outcome of hormonal ovarian superstimulation in small ruminants, and the total dose of exogenous gonadotropins used may be one of the causes of this condition. There were two major objectives of the present study: (1) to evaluate the effects of different superovulatory doses of porcine follicle-stimulating hormone (pFSH) on the biometry, blood perfusion (Doppler), and echotextural characteristics of luteal structures; and, (2) to determine the usefulness of biometric, vascular, and echotextural luteal variables, as well as measurements of circulating progesterone (P4) concentrations for early detection of PRCL in superovulated Santa Inês ewes. Twenty-seven Santa Inês ewes received an intravaginal P4-releasing device (CIDR) from Days 0 to 8 (Day 0 = random day of the anovulatory period). An IM injection of d-cloprostenol (37.5 µg) was given at the time of the CIDR insertion and withdrawal. On Day 6, all the ewes received 300 IU of eCG IM and were divided into three treatment groups (each n = 9): G100 (100 mg); G133 (133 mg); and G200 (200 mg of pFSH) administered IM every 12 h in eight injections. Transrectal ovarian ultrasonography and jugular blood sampling for serum P4 measurements were performed on Days 11 to 15. On the day of embryo recovery (Day 15), all the ewes underwent diagnostic videolaparoscopy and were classified, based on their luteal characteristics, into three response groups: nCL (ewes with normal CL only); rCL (ewes with regressing CL only); and ewes with both nCL and rCL following the superovulatory regimen. Our present results indicate that the total pFSH doses of 100 mg and 200 mg result in similar ovulatory responses and luteal function/biometrics, although the percentage of donor ewes with nCL was greater (p < 0.05) for G100 compared with the G200 animals. An application of 133 mg of pFSH was associated with diminished luteogenesis. Lastly, circulating P4 concentrations, ultrasonographic estimates of total luteal area, and CL pixel heterogeneity (standard deviation of numerical pixel values) are promising markers of luteal inadequacy in superovulated ewes.

Luteal function in cyclic goats treated with human chorionic gonadotropin administered by intramuscular or intravaginal routes at the time of artificial insemination.

Human chorionic gonadotropin (hCG) has been used to improve goats reproductive efficiency. This study aimed to (i) evaluate if hCG administered by the intramuscular (i.m.) or intravaginal (i.vag.) route can be detected by a rapid ß-hCG test in blood plasma samples and (ii) document ovarian effects of hCG administered by both routes at the time of artificial insemination (AI) performed 60 h after oestrus synchronization in goats. Twenty-two Alpine goats received two i.m. injections of 30 µg of d-cloprostenol (Prolise®, Tecnopec, São Paulo, Brazil) 7.5 days apart. One day after the onset of oestrus (at the time of AI), the goats were randomly allocated to one of the three groups that received: control (n = 7): 0.3 ml of saline solution intravaginally; hCGi.m. (n = 7): 300 IU of hCG (Vetecor®; Hertape-Calier, São Paulo, Brazil) i.m. and hCGi.vag. (n = 8): 300 IU of hCG deposited intravaginally. Blood samples were drawn at -1, 3, 6, 9 and 24 h after as well as on days 3, 7, 10, 13, 17 and 21 after hCG treatment/AI. All animals tested negative for hCG (ECO Diagnóstica, Corinto, Brazil) at -1 h, and all control animals tested negative throughout the entire blood collection period. All hCGi.m. animals tested positive from 3 h until D3 post-AI but only 50% of hCGi.vag. goats tested positive during the present study. In all animals studied, mean circulating P4 concentrations increased (p < .05) from D3 to D7 after AI and then declined (p < .05) from D10 to D17 in control and hCGi.m. groups and from D17 to D21 in the hCGi.vag. group. Total cross-sectional luteal area (CLA), mean colour Doppler area (DA), DA/CLA, mean high-velocity Doppler area and HVDA/CLA all declined (p < .05) by D17-D21 in all animals studied. In summary: (i) human chorionic gonadotropin could consistently be detected in blood samples using the rapid ß-hCG test only in the hCGi.m. group; and (ii) there were no significant differences in the mean pregnancy rate, circulating P4 concentrations and various luteal parameters studied among Control, hCGi.m. and hCGi.vag. dose.

Hormonal-Induced Cervical Relaxation During Diestrus in Ewes: Cervical Transposing Feasibility and Use of hCG for Rescuing Disrupted Luteal Function.

Cervical relaxation (CR) was performed in ewes during diestrus, to prospect a feasible protocol for non-surgical embryo transfer (NSET). In Trial 1, naturally mated ewes (n=13) received CR protocols with estradiol benzoate (EB, 1 mg on D6) and oxytocin (OT, 50 IU on D7) only (G-EB+OT) or associated with human chorionic gonadotropin (hCG, 300 IU on D7, G-EB+OT+hCG) and were compared to non-hormonally treated (G-control) ewes. Estradiol concentration increased (P<0.05) from D6 to D7 in G-EB+OT and G-EB+OT+hCG, then decreased (P<0.05) on D8. The G-EB+OT had reduced (P<0.05) progesterone on D8, and 0% (0/5) pregnancy, while G-control had 100% (3/3) conception rate. The treatment with hCG mitigated the progesterone drop on D8 and resulted in 50% (2/4) conception rate. In Trial 2, cervical transposing success after treatment with EB, OT, and hCG on D6 and D7 was assessed in nulliparous (n=25) and multiparous (n=30) ewes. The EB+OT+hCG protocol allowed efficient cervical transposition (~78% of cervical transposition success into ~5 min) in nulliparous and multiparous females and did not alter total and blood perfusion areas of corpora lutea (CL). The use of EB+OT+hCG was efficient to ensure effective cervical transposing in nulliparous and multiparous ewes, with no apparent deleterious effect on CL function, and may contribute to animal welfare, allowing a less invasive NSET procedure.

Study of the factors affecting embryo yields and quality in superovulated Morada Nova ewes that underwent non-surgical uterine flushing.

The present study compared the outcomes of in vivo embryo production in Morada Nova ewes subjected to either 9-day (G-9SOV , n = 21) or 12-day (G-12SOV , n = 21) progesterone (P4 )-based estruses synchronization protocol coupled with superovulatory treatment with decreasing doses of porcine follicle-stimulating hormone (133 mg of pFSH given over 3 days). Non-surgical embryo recovery (NSER) was performed 6-7 days after the onset of oestrus. Total antral follicle count doubled from the first to the sixth pFSH dose in both groups (p < .05). Oestrus responses did not vary between the two groups of animals (95.2%). Corpora lutea (CL) were detected in 85.0% and 60.0% of ewes that previously manifested oestrus behaviour in G-9SOV and G-12SOV respectively. NSER was successfully completed in 86.2% of ewes that had CL (p > .05). The mean number of CL per ewe/successfully flushed donor ewe was greater (p < .05) in G-12SOV (12.3 ± 1.7/12.1 ± 1.9) than in G-9SOV (7.9 ± 1.4/8.2 ± 1.6). Mean numbers of retrieved blastocysts and viable embryos were greater (p > .05) in G-12SOV (5.8 ± 1.9 and 3.7 ± 1.7) than G-9SOV (3.5 ± 1.1 and 0.8 ± 0.3 respectively). The total follicle count (all follicles ≥2 mm in diameter) at the sixth pFSH dose (at P4 -device removal) was positively correlated (p < .05) with the number of CL (r = .95) and viable embryos (r = .91) in G-12SOV . The ewes with ≥10 Cl (48% of all flushed donors) yielded 80.5% of viable embryos. In summary: (a) Morada Nova ewes from G-12SOV group had better superovulatory responses compared with G-9SOV group; (b) total follicle count at the last pFSH dose was a good predictor of superovulatory responses only in the ewes primed with P4 for 12 days; and (c) animals with ≥10 ovulations are main contributors to viable embryo production in Morada Nova ewes.

Human chorionic gonadotropin affects original (ovulatory) and induced (accessory) corpora lutea, progesterone concentrations, and pregnancy rates in anestrous dairy goats.

Two experiments were conducted in acyclic Alpine (A) and Saanen (S) goats that received intravaginal sponges containing 60 mg of medroxyprogesterone acetate for 6 days, as well as 200 IU of eCG and 30 µg d-cloprostenol i.m. 24 h before sponge removal. On day 7 (day 0 = onset of synchronized estrus), all goats were randomly divided into two groups: animals treated with 300 IU of hCG i.m. (hCG; Exp.1: n = 8A; Exp.2: n = 75A + S) and untreated controls (Control; Exp.1: n = 8A; Exp. 2: n = 70A + S). In Exp.2, all goats were artificially inseminated. Transrectal ovarian ultrasonography and blood collection were done on days 7, 10, 13, 17, and 21 (Exp.1), and pregnancy detection on day 60 (Exp.2). Estrus and ovulations occurred in five hCG and seven Control animals. Accessory CL (aCL) were detected in all hCG does. The total luteal area of ovulatory corpora lutea (oCL) increased (P < 0.05) on day 10 in hCG does and remained greater (P < 0.05) than in Control until day 21. Total and high-velocity color Doppler area were greater (P < 0.05) for oCL of hCG does on days 13 and 17. Progesterone concentrations were greater (P < 0.05) in hCG does from days 13 to 21 and related directly to the total luteal and oCL area for the duration of the study in all does. The pregnancy rate was higher (P < 0.05) in hCG than in Control by 22.5 %. Human chorionic gonadotropin given on day 7 of the synchronized estrous cycle positively affected CL function and pregnancy rates in seasonally anovular dairy goats.

Overnutrition of Ewe in Late Gestation and the Impact on Placental Efficiency and Lamb's Performance.

The objective of the study was to evaluate energy or energy/protein supplementation in the ewe diet, in the last third of gestation, on maternal placental and endocrine characteristics, as well as its effects on the behavior of neonatal lambs and productive performance until weaning. A total of 128 ewes were used, and the experimental diet was fed from 100 days gestation until lambing, with the birth of 172 lambs. The ewes were distributed into three treatments: CTL (control, n = 43) with metabolizable energy (ME) and crude protein (CP) intake according to NRC (1985); ME (energy supplementation, n = 44) plus 21% ME; and MECP (energy/protein supplementation, n = 41) plus 26% ME and CP. Body weight, body condition score, serum hormone concentrations, placental characteristics, lamb performance and behavior, and production efficiency of the ewe from lambing to weaning were measured. ME and MECP ewes were heavier before (p = 0.006) and just after lambing (p = 0.002) and had higher serum triiodothyronine (p = 0.001) and cortisol (p = 0.004) concentrations on the day of lambing. ME ewes had higher placental efficiency (p = 0.036) and lower total cotyledon weight (p = 0.011). ME and MECP diets increased both lamb birth weight (p = 0.015) and weaning weight (p = 0.009). Production efficiency at birth and at weaning was not influenced (p > 0.05) by treatments. Lamb behavior was influenced by the ME and MECP diets, reducing the time to kneel (p ≤ 0.05), to try to stand (p ≤ 0.05), and the latency to stand (p ≤ 0.005). It is concluded that overnutrition in the last third of gestation was positive for the ovine production system, with improved production rates, hormonal profile, placental characteristics, and neonatal behavior.

Assessment of dopperfluxometric indices of maternal-fetal structures in pregnant ewes.

The aim of this study was to evaluate the blood flow of the uterine artery, fetal aorta and umbilical artery in the physiological pregnancy of sheep by means of pulsed Doppler throughout the gestational period. Thirty Santa Inês ewes weighing between 45.4±4.3 kg and aged 2 to 5 years were selected. The evaluations were carried out weekly from the 3rd to the 21st gestational week. Peak systolic velocity (PSV), end diastolic velocity (EDV) and resistance index (RI) were obtained. Analysis of variance was performed, and the minimum significant comparison of means was obtained by the BH test with adjusted P<0.05. The results were expressed as mean ± standard error. For the fetal aorta, there was an increase in the EDV values and a decrease in the PSV and RI throughout pregnancy. For the uterine artery, PSV and EDV did not present significant variation, whereas the RI showed a reduction in the last week. Increased EDV values were found for the umbilical artery throughout pregnancy. For the PSV there was no significant difference, as the RI was reduced at the end of pregnancy. The results obtained are expected to contribute to a more complete understanding of the hemodynamic changes resulting from pregnancy.

Effect of eCG in a short-term synchronization treatment on ovarian status, estrus synchrony, and ovulation in dairy goats managed under tropical conditions.

The aim of this study was to assess the need of using eCG on short-term estrus synchronization protocol in nulliparous (NUL) and multiparous (MULT) dairy goats during the breeding season. Alpine (n = 20), Nubian (n = 20), and Saanen (n = 16) goats received 60 mg medroxyprogesterone acetate intravaginal sponges for 6 days plus 30 µg d-cloprostenol and 200 IU eCG (G-eCG, n = 28) or saline (G-Control, n = 28) 24 h before sponge removal. The NUL and MULT goats of each breed were equally assigned into the two treatments. Transrectal ultrasonography was used to evaluate ovulatory parameters, and teaser goats were used for estrus detection every 12 h from sponge removal to ovulation. eCG did not affect (P > 0.05) estrus response (~86%), diameter of ovulatory follicles (~6.8 mm), and number of ovulations (~1.6). Nevertheless, eCG led to earlier (P < 0.05) ovulation (G-eCG = 65.1 and G-Control = 73.2 h) and increased (P < 0.05) the ovulation rate (G-eCG = 96.4% and G-Control = 67.9%). In the absence of eCG, no differences regarding reproductive parameters (P > 0.05) were found between parity orders. Alpine MULT goats underwent a superior (P < 0.05) number of ovulations (2.2) in comparison to NUL goats (1.3). In conclusion, the exclusion of eCG from short-term estrus synchronization protocol did not interfere with estrus response but decreased the ovulation rate.

Vaginal cytology and cervical mucus as tools to predict ovulation time in small ruminants.

The possibility of using cervical mucus and vaginal cytology as tools to predict ovulation time was assessed in 11 ewes and 11 does raised under tropical conditions. Every 12 h from progesterone removal to ovulation, estrus behavior, cervical mucus, vaginal cytology, and ovarian ultrasound exams were performed. In goats, vaginal cytology had 88% of accuracy on detecting the ovulation time. However, in sheep, there was no cell pattern in the vaginal cytology and cervical mucus varied at ovulation. In conclusion, both vaginal cytology and mucus evaluation may be useful tools to determine the ovulation time in goats; however, both strategies are less accurate in sheep.

NonSurgical Embryo Recovery from Estrus-Synchronized or Superovulated Morada Nova Ewes: A Feasible Strategy for Sheep Embryo Banking.

This study assessed the feasibility of in vivo embryo production and nonsurgical embryo recovery (NSER) in Morada Nova ewes (an endangered native Brazilian breed of sheep) subjected to different estrus synchronization and/or superovulation protocols. Ewes received intravaginal sponges soaked with 60 mg medroxyprogesterone acetate (MAP), which were kept in place for six (G6; n = 12), nine (G9; n = 12), or 12 (G12; n = 12) days. Half of the ewes in each group remained estrus synchronized only (SYNCH) and the other half was superovulated (SOV) with 133 mg porcine follicle-stimulating hormone (pFSH). There were no differences (p > 0.05) in antral follicle counts determined with ultrasonography 60 hours before MAP sponge removal (or at the time of the first pFSH dose) among G6 (6.4 ± 0.9), G9 (6.2 ± 0.7), and G12 (5.5 ± 0.6). Estrus responses and NSER success rates did not vary (p > 0.05) among the three progestin-treatment groups of ewes for either estrus-induced or superovulated animals. The onset of estrus occurred 10-12 hours later (p < 0.01) in G9SYNCH ewes compared with G6SYNCH and G12SYNCH, and the duration of estrus was ∼19 hours greater (p < 0.01) in G9SOV than in G6SOV. The average duration of the NSER procedure was 32.6 ± 1.3 minutes. At least one structure was recovered in 85.7% of synchronized and in 87.5% of superovulated ewes. Viable embryo recovery rates were also similar (p > 0.05) for G6 (1.0 ± 0.3 and 2.5 ± 1.5), G9 (1.3 ± 0.5 and 4.8 ± 2.0), and G12 groups (1.0 ± 0.3 and 4.8 ± 2.3; estrus-synchronized and superovulated ewes, respectively). In conclusion, progestogen pretreatment of different durations and NSER can be employed in Morada Nova ewes, resulting in reasonable viable embryo recovery rates in both estrus-synchronized and superovulated animals. Therefore, both techniques are suitable for use in commercial settings as well as small ruminant conservation programs.

Luteotropic effects of human chorionic gonadotropin administered 7.5 days after synchronous estrous induction in Morada Nova ewes.

This study was conducted in ewes to assess effects of human chorionic gonadotropin (hCG) administration after imposing an estrous induction treatment regimen. Ewes (n = 115) were treated with a 60 mg medroxyprogesterone-intravaginal-sponge for 6 d plus 200 IU of equine chorionic gonadotropin (eCG) im and 37.5 µg d-cloprostenol im 36 h before sponge removal (Day 0). After natural mating, ewes having at least one corpus luteum (CL; n = 108) were administered either 1 mL of saline (G-Control; n = 53) or 300 IU of hCG (G-hCG; n = 55) on Day 7.5 after sponge removal (Day 0). Ovarian ultrasonography and blood collection were performed on Days 7.5, 13.5, 17.5, 21.5, and 30.5. Accessory CL (aCL) were observed in 81.5 % (G-hCG) and 0.0 % (G-Control) of ewes (P = 0.0001). Diameter, area, and volume of luteal tissue were greater (P < 0.05) in G-hCG from Day 13.5 to 30.5. Progesterone (P4) concentrations were greater (P < 0.05) on Days 13.5, 17.5, 21.5 and 30.5 for ewes of the G-hCG group. Pregnancy percentage was similar (P = 0.25) between groups [47.1 % (G-control) compared with 60.0 % (G-hCG)], although total number of lambs produced by estrous synchronized ewes was greater (P = 0.005) in ewes of the G-hCG group (90.9 % compared with 66.0 %). In conclusion, hCG administration 7.5 days after sponge removal from Morada Nova ewes during the non-breeding season is an effective treatment to induce aCL formation, improve luteal tissue biometry and P4 concentrations, and to enhance the total number of lambs born.

In vivo embryo production and recovery in lacaune ewes after imposing a superovulation treatment regimen is related to pFSH dose.

This study was conducted to assess effects of different doses of pFSH on follicular recruitment, superovulatory response, ova/embryo recovery, and embryo yield in lactating ewes. Ewes (n = 24) had a superovulation treatment regimen imposed. All ewes were implanted with a progesterone intravaginal device for 9 d, and administered either 100 (G-100) or 200 (G-200) mg pFSH, proportioned into six doses administered at 12-h intervals, starting 60 h before device removal. At 7 days subsequent to progesterone device removal, there were non-surgical embryo recoveries (NSER) from ewes having three or more corpora lutea. At the time of the first pFSH injection, number of antral follicles were similar (P < 0.05) between ewes in the G-100 and G-200 group, however, there were more 3.1-4.0 mm follicles in ewes of the G-200 than G-100 group at the time of the second pFSH administration. Estrous response and CL number were less (P < 0.05) in ewes of the G-100 (66.7 % and 2.6 ±â€¯0.7) than G-200 (91.7 % and 11.6 ±â€¯1.2) group. There were embryo collections from 100 % and 90.9 % of ewes in the G-100 and G-200 groups, respectively (P > 0.05). Viable embryo numbers and ova/embryo recovery rate were greater (P < 0.05) in ewes of the G-200 (6.9 ±â€¯1.1 and 67.8 %) than G-100 (1.0 ±â€¯0.5 and 27.6 %) group. A dose of 200 mg pFSH was more effective in inducing a superovulatory response and embryo yield after NSER in ewes, however, the 100 mg dose was insufficient for these purposes.