Intravaginal Sponges Research Articles

Medroxyprogesterone acetate or long-acting progesterone in the biostimulation of lambs

The aim of this study was to evaluate the response of prepubertal ewe lambs to exogenous administration of either medroxyprogesterone acetate (MAP) or long-acting progesterone (LAP) together with biostimulation. Two Pool Dorset adult males and 75 mixed-breed prepubertal ewe lambs (average of 179 days-old and 30.0kg) were used. The females were randomly assigned to three different groups. In the first group the females were submitted to the insertion of intravaginal sponges containing MAP (60 mg) for 12 days and were then biostimulated for eight weeks. In the second group the females were submitted to a single injection of LAP (225 mg) and then to biostimulation for eight weeks. In the last group, the females were only submitted to biostimulation for eight weeks. Animals were considered cyclic when plasma progesterone (P4) concentration exceeded 1.0 ng/mL in at least one of two consecutive blood samples taken within a 7-day interval in three distinct experimental moments. After treatments 93.3% of the females disregarding their group started their cyclicity and most of them (92.0%), continued to be cyclic after 63 days of either MAP or LAP together with biostimulation under both male and female effect. We conclude that prepubertal ewe lambs when submitted to protocols of either MAP or LAP followed by biostimulation result in puberty at the 7 month of age. It can be deducted that some ewe lambs submitted to the administration of either MAP or LAP together with biostimulation promoted a multiplier effect upon the other young females that were then stimulated to start cyclicity.

Accelerated lambing achieved by a photoperiod regimen consisting of alternating 4-month sequences of long and short days applied year-round1

The objective of this work was to evaluate the reproductive performance of ewes exposed to a photoperiodic regimen consisting of continuous alternating 4-mo periods of long days (LD: 16 h of light/d) and short days (SHD: 8 h of light/d) in an accelerated lambing program of 3 lambings in 2 yr. A total of 211 prolific Rideau Arcott ewes were assigned to the photoperiodic treatment, whereas 37 ewes were maintained under natural annual variation in day length (control group). Ewes under the photoperiod regimen were divided into 4 subgroups (A, B, C, D). All these groups of ewes were exposed to the same light regimen, but the LD and SHD light sequences were staggered by 2 mo to permit the evaluation of the effect of time and season of mating on performance of the ewes treated with the photoperiod. The control ewes were treated with intravaginal sponges in the out-of-season breeding periods (conventional approach). Each group of ewes was studied over 3 reproductive cycles. Two groups of rams exposed to alternating 2-mo sequences of LD and SHD were used for mating. The short mean interval between ram introduction and conception for the groups exposed to artificial photoperiod (9.4 d) confirmed the effectiveness of the treatment to induce intense sexual activity. For the 12 breeding periods studied (8 in out-of-season and 4 in sexual season), fertility rate of the ewes treated with photoperiod, mated at various times of year, was 91.6%, which is comparable with the fertility normally seen in the natural breeding season. The number of lambs born/ewe remained constant across reproductive cycles and was greater in photoperiod-treated groups (2.81 vs. 2.27 for photoperiod and control groups, respectively; P = 0.0002). Groups exposed to photoperiod treatment obtained better fertility rate than the control group in out-of-season breeding (91.1 vs. 76.3%; P = 0.016). Ewes managed under the photoperiod regimen produced 1.38 lambings/yr and 69% of them lambed 3 times in 2 yr. Overall, the ewes in the photoperiodic treatment produced annually 3.78lambs/ewe. The reproductive performances achieved throughout the years indicate that the photoperiodic program tested, consisting of continuous alternating 4-mo periods of LD and SHD, allows control of the annual reproductive cycles in ewes.

SINCRONIZAÇÃO DE ESTRO COM PROSTAGLANDINA F2? versus PROGESTÁGENO ASSOCIADO À GONADOTROFINA CORIÔNICA EQUINA (eCG) EM OVELHAS SANTA INÊS NO DISTRITO FEDERAL, BRASIL

The aim of this study was to compare two protocols of estrus synchronization in Santa Inês ewes. Thirty-eight ewes were randomly divided into two groups of estrus synchronization: protocol PGF2? (two doses of 0.530 mg of PGF2?, nine days apart) and protocol MAP+eCG (intravaginal sponge impregnated with medroxyprogesterone acetate, for 12 days, and then an injection of 250 IU of eCG). The experiment was in a cross-over design, two estrous cycles apart. On the final day of protocol, a transrectal ultrasound examination was carried out to measure the size of the largest and second largest ovarian follicles and on day 7 of estrous cycle blood was collected to measure serum P4 concentration. Laparoscopy was carried out on day 11 after the end of protocols to count corpora lutea. Synchronization rate, size of largest and second largest ovarian follicles, interval between the end of the protocol to estrus and ovulation rate did not differ between protocols. Ewes synchronized with MAP+eCG had greater serum P4 concentrations than ewes synchronized with PGF2? (3.9 and 2.8 ng/mL, respectively, P

Characterization of Oestrous Induction Response, Oestrous Duration, Fecundity and Fertility in Awassi Ewes During the Non‐breeding Season Utilizing both CIDR and Intravaginal Sponge Treatments

The aim of this study was to investigate characterization of oestrous response, onset of induced oestrus, oestrous duration, fecundity and fertility in Awassi ewes treatment with intravaginal sponges and Controlled Intravaginal Drug Release (CIDR) devices in combination with pregnant mare serum gonadotropin (PMSG) under local environmental conditions during the non-breeding season. A total of 62 ewes were divided into three groups. Group CIDR (n = 20) was treated with CIDR devices for 12 days and 400 IU PMSG was injected upon removal of the CIDR. For ewes in Group Sponge (SP) (n = 24), 30 mg fluorogestone acetate was administered to the sheep for 12 days and 400 IU PMSG was injected upon withdrawal of the sponge. Group Control (CON) (n = 18) served as a control group and received no treatment. Adult, intact and sexually experienced Awassi rams were introduced to all groups at the time when the intravaginal devices were removed. There were no significant differences in terms of oestrous response (CIDR: 90%, SP: 87.5%), time to onset of oestrus and duration of induced oestrus between the CIDR and SP groups. The oestrous response of treatment groups was significantly greater (p < 0.05) than in the control ewes. There were no significant differences in pregnancy (CIDR: 70%, SP: 70.8%), lambing (CIDR: 85%, SP: 79.2%) and fecundity rates between ewes treated with CIDR and those treated with sponges. However, pregnancy and lambing rates were significantly (p < 0.05) higher in ewes treated with CIDR or sponges when compared with those in the control group. It was concluded that it is possible to induce fertile oestrus, successful pregnancy and lambing with the treatment of either CIDR or intravaginal sponge in combination with PMSG in Awassi ewes during the non-breeding season.

Efficiency of methods applied for goat estrous synchronization in subtropical monsoonal climate zone of Southwest China

Two experiments were undertaken to select the efficient method applied for goat estrous synchronization. In experiment 1, a total of 120 does (Capra hircus) were divided into five groups with a randomized block design, and the does of treatment 1 were synchronized by intravaginal sponges impregnated with 30 mg Levonorgestrel inserted for 10 days. Does of treatments 2, 3, and 4 were treated with further injection of 25 IU follicle-stimulating hormone (FSH), 0.05 mg prostaglandin F2 alpha (PGF(2alpha)), and 25 IU FSH + 0.05 mg PGF(2alpha) at sponge withdrawal, respectively. The does in the control group (n = 40) without estrous synchronization treatment and natural estrous does were observed. In experiment 2, a total of 140 does in five goat farms in breeding and non-breeding seasons were treated with the selective efficient procedure. The results presented that all the employed treatments were capable of inducing and synchronizing estrous goats. According to estrous response and economy, the use of intravaginal sponges impregnated with 30 mg Levonorgestrel and 0.05 mg PGF(2alpha) (treatment 3) is the first choice for estrous synchronization, and 95.0% of synchronized does demonstrated estrus, which was significantly higher than that of treatment 1 (P < 0.05) and control group (P < 0.01). The percentages of ovulating of treatments 3 and 4 were the same (95.0%), which were significantly higher than that of treatment 1 (P < 0.01). The ovulation rates among different groups were not significant (P > 0.05). When the selective procedure was applied to five goat farms, 85.7% (120/140) of does demonstrated estrus, and the kidding percentage, litter size, and prolificacy rate were 53.6%, 0.95, and 177%, respectively.

Morphological features and vascularization study of caprine cyclic corpus luteum

Corpus luteum is a temporary endocrine gland that regulates either the estrous cycle and pregnancy. It presents extreme dependency on the adequate blood supply. This work aims to evaluate goat corpus luteum (CL) vascular density (VD) over the estrous cycle. For that purpose, 20 females were submitted to estrus synchronization/ovulation treatment using a medroxyprogesterone intra-vaginal sponge as well as intramuscular (IM) application of cloprostenol and equine chorionic gonadotrophine (eCG). After sponge removal, estrus was identified at about 72hs. Once treatment was over, female goats were then subdivided into 4 groups (n=5 each) and slaughtered on days 2, 12, 16 and 22 after ovulation (p.o). Ovaries were collected, withdrawn and weighted. CL and ovaries had size and area recorded. Blood samples were collected and the plasma progesterone (P4) was measured through RIA commercial kits. The VD was 24.42±6.66, 36.26±5.61, 8.59±2.2 and 3.97±1.12 vessels/mm² for days 2, 12, 16 and 22 p.o, respectively. Progesterone plasma concentrations were 0.49±0.08, 2.63±0.66, 0.61±0.14 and 0.22±0.04ng/ml for days 2, 12, 16 e 22 p.o, respectively. Studied parameters were affected by the estrous cycle phase. Values greater than 12 p.o were observed. In the present work we observed that ovulation occurred predominantly in the right ovary (70% of the animals), which in turn presented bigger measures than the contra lateral one. There is a meaningful relationship between the weight and size of the ovary and these of CL (r=0.87, r=0.70, respectively, p&lt;0.05). It is possible to conclude that morphology of goat's ovaries and plasma progesterone concentration changed according to estrous cycle stages. We propose these parameters can be used as indicators of CL functional activity.

Repeated administration of hCG on follicular and luteal characteristics and serum progesterone concentrations in eCG-superovulated does

The objective of this study was to evaluate the effect of repeated administration of hCG on the follicular and luteal charactristics and serum progesterone concentrations in eCG-superovulated Merghoz does. The estrous cycles of 40 adult does were synchronized following a 12-day treatment with intravaginal sponges (Chronogest ®), with 1000 IU eCG injected 2 days prior to sponge removal. On the day of sponge removal (day 0), the females were randomly assigned to 4 treatments. The first group (h1; n = 10), received an i.m. hCG injection (500 IU; Pregnyl ®) on the day 0. The second group (h2; n = 10), received an i.m. hCG (500 IU) injection on days 0 and 1, while the third group (h3; n = 10) received i.m. hCG (500 IU) on days 0–2. Does in the control group ( n = 10) did not receive any hCG treatment. Blood samples were taken on days 0, 1, 2, 5 and 8 and analysed for serum progesterone (P4) concentrations. Premature luteal regression was considered to have occurred if the P4 concentrations declined to less than 1 ng/mL by day 8 following sponge removal. On day 8 goats were slaughtered and ovaries recovered for evaluation. In each case the CL's were counted and classified according to their appearance – as normal or as regressing. According to the serum P4 concentration on day 8, 42.8% (3/7), 0% (0/8), 0% (0/7) and 0% (0/6) of the goats underwent premature luteal regression in the control, h1, h2 and h3 groups, respectively. The control group was the only group in which there was a significant increase in the number of regressing CL's on day 8 (5.1 ± 1.5; P < 0.05). The characteristics of the CL's, including the mean diameter and weight was greater ( P < 0.05) in the h2 and h3 groups, compared to the control group (10.9 ± 0.7, 11.0 ± 0.9 vs. 7.9 ± 1.1 mm and 640.0 ± 101.4, 691.7 ± 111.7 vs. 294.3 ± 143.7 mg, respectively). No differences ( P < 0.05) in the number of large follicles present on the ovaries were recorded on day 8 between the groups. However, the diameter of the large follicles present on the ovaries on day 8 was greater ( P < 0.05) in goats in the h2 and the h3 groups, compared to the control (15.3 ± 0.6 and 12.9 ± 0.3 vs. 8.6 ± 1.7 mm, respectively; P < 0.05). The serum P4 concentrations were higher in the h2 and the h3 groups than in the control group on days 2 and 8. The serum P4 concentrations on day 8 also recorded a positive correlation with the normal CL's, CL diameter and CL weight ( P < 0.001). These results indicate that the repeated administration of hCG post-sponge removal presumably prevents premature luteal regression and improves luteal function in goats after eCG-superovulatory treatment. However, neither the number of persistent follicles nor the diameter of the persistent follicles was reduced.

Effect of estradiol benzoate used at the start of a progestagen treatment on superovulatory response and embryo yield in lactating and non-lactating llamas

Two experiments were conducted to determine the effect of estradiol benzoate (EB) and intravaginal progestagen treatment on ovarian follicular dynamics and superstimulatory response in eCG-treated llamas. The purpose of Experiment 1 was to evaluate the effect of EB and progestagen treatment starting at different phases of dominant follicle (DF1) development on regression pattern and subsequent follicle wave emergence (WE2) in lactating and non-lactating llamas. Early lactating ( n = 24, 30 ± 4 days postpartum) and non-lactating ( n = 24) females were assigned in equal numbers ( n = 8) to one of three groups according to the phase of DF1 (growing, static or regressing) determined by ultrasonography from day −3 to day 0. At day 0, llamas received an intravaginal sponge (MPA, 150 mg) and 5 mg of MPA (i.m.). Half of the females ( n = 4) of each group were injected with 2 mg (i.m.) of EB and half were not (control group). A 2 × 2 × 3 (lactational status, EB treatment and follicular phases) factorial design was used. Each sponge was removed 8 days later. Ovaries were monitored from day 0 to day 12. Daily blood samples were taken to determine 17β-estradiol (E 2) profiles from day 0 to day 8. The DF1 regression pattern was not affected ( P > 0.05) by the phase of follicle wave at the start of the treatment or any interactions among main effects. Follicle wave emergence in EB-treated llamas was delayed ( P < 0.05) by 2.3 days compared with non-treated llamas. Following EB treatment, plasma concentrations of E 2 were greater ( P < 0.05) from day 1 to day 5 in the treated than in non-treated females, but not from day 6 onward ( P > 0.05). Experiment 2 was designed to evaluate the effect of this treatment on the ovarian superovulatory response and embryo yield following eCG treatment administered on day of follicular wave emergence as determined in the Experiment 1. The same lactating ( n = 18, 61 ± 4 days postpartum) and non-lactating ( n = 18) llamas at random stages of follicle wave were treated as those in Experiment 1 and received 1200 IU of eCG at the time of WE2 (EB-treated = day 6.5 and non-treated = day 4.5). Llamas were mated 5 days after sponge withdrawal. A second mating was allowed 24 h later. Embryos were collected between 7 and 8 days after the first mating and blood samples were taken to determine progesterone (P 4) concentrations. The mean number of follicles on day of mating and the number of CL on day of embryo collection were not affected by lactational status, EB treatment or their interactions ( P > 0.05). Ovulation rate and mean (±SEM) number of recovered embryos for EB treatment group (67.5% and 2.4 ± 0.4) were greater ( P < 0.05) than for no EB treatment (51.1% and 1.1 ± 0.4). Plasma P 4 concentrations and number of CL per llama were correlated ( r = 0.49; P = 0.014). In conclusion, progestagen plus EB treatment facilitates the prediction of the emergence of a new follicular wave approximately 6 days after treatment and resulted in a higher ovulation rate and embryo production in ovarian superstimulated llamas regardless of lactational status.

Reproductive Characteristics of Awassi Ewes Under Cornell Alternate Month Accelerated Lambing System

We investigated the reproductive responses of Awassi ewes under Cornell alternate month accelerated lambing (CAMAL) system. Ewes were randomly allocated to two experimental groups. The first group (control group) consisted of 20 ewes exposed to rams in September under conventional management system, while the second group (CAMAL) was divided into four sub-flocks containing each 20 ewes exposed to rams to obtain three lambing in two years with different breeding and lambing months. In CAMAL ewes, oestrus was synchronized using intra-vaginal sponges with progesterone and Pregnant Mare Serum Gonadotropin (PMSG) administration. The results obtained revealed that within CAMAL group, the percentage of animals in heat, onset of oestrus, litter size, birth weight and weaning weight were affected by mating months. September and November were the most appropriate months for oestrus ratio (97.5%) and litter size (1.18 and 0.98, respectively). Lambs of control group were heavier at birth and weaning than those of CAMAL group. On the other hand lamb yield was not affected statistically by the CAMAL administration.

Oestrus synchronization with short-term and long-term progestagen treatments in goats: the use of GnRH prior to short-term progestagen treatment

The aim of the present study was to determine the efficacy of the synchronization of oestrus using short- and long-term progestagen treatments in Hair goats at the onset of the breeding season, and to evaluate the effect of the exogenous GnRH administration immediately prior to short-term progestagen treatment on the reproductive performance. A total of 75 Hair goats, aged 2.5–5 years-old were used in this experiment. Goats were divided equally into three groups (n=25 per group). Animals in LT-FGA (long-term progestagen treatment), ST-FGA (short-term progestagen treatment) and Gn-ST-FGA (GnRH-short-term progestagen treatment) groups received an intravaginal sponge (day 0) containing 30 mg fluorogestone acetate (FGA) for 14, 8 and 8 days, respectively, plus 75 µg cloprostenol i.m. 24 h before sponge removal and 400 I.U. equine chorionic gonadotrophin (eCG) i.m. at the time of sponge removal. In addition, the goats in the Gn-ST-FGA group received 10.5 µg busereline acetate i.m. at the time of sponge insertion (day 0). Oestrus response for all treatment groups was 100%. The mean interval from sponge removal and the onset of oestrus for the LT-FGA, ST-FGA and Gn-ST-FGA groups was 28.0±1.0 h, 28.8±1.1 h and 33.1±2.0 h, respectively. No significant difference in onset of oestrus among groups was recorded. The pregnancy rate, kidding rate, multiple kidding rates and litter size were 72.0, 61.1, 45.5% and 1.6 in the LT-FGA, 70.8, 76.5, 69.2% and 1.8 in the ST-FGA and 58.3, 78.6, 63.6% and 1.6 in the Gn-ST-FGA groups, respectively. The pregnancy rates were similar in the LT-FGA (72.0%) and ST-FGA (70.8%). However, the kidding rate, multiple kidding rates and litter size were numerically higher in the ST-FGA (76.5%, 69.2% and 1.8, respectively) group than in the LT-FGA (61.1%, 45.5% and 1.6, respectively) group. Although not statistically different, pregnancy rate and litter size was lower in the Gn-ST-FGA group (58.3% and 1.6, respectively) compared with the ST-FGA (70.8% and 1.8, respectively).In conclusion, it can be said that oestrus synhcronization with long-term progestagen treatment (14 d-FGA- PGF2α- eCG) could be replaced with short-term progestagen treatment (8 d-FGA-PGF2α- eCG) without a reduction in oestrus rate and fertility parameters in lactating goats at the beginning of breeding season. However, the use of GnRH prior to short-term progestagen treatment (8 d-FGA-PGF2α- eCG) do not have a positive effect on fertility parameters in goats.

Fertility Rates of Ewes Treated with Medroxyprogesterone and Injected with Equine Chorionic Gonadotropin plus Human Chorionic Gonadotropin in Anoestrous Season

The aim of the present paper was to investigate the efficiency of the equine chorionic gonadotropin (eCG) plus human chorionic gonadotropin (hCG) associated with medroxyprogesterone acetate (MAP) to estrous ewes synchronization. Ninety Texel ewes were investigated during seasonal anoestrous. The ewes received intravaginal sponges containing MAP (60 mg) for nine days. At the time of sponges' withdrawal, the ewes were divided into three groups (G): (1) receiving 2 mL of saline i.m. (n = 30), (2) receiving eCG 400 IU i.m. (n = 30), and (3) receiving eCG 400 IU plus hCG 200 IU i.m. (n = 30). Twelve h after sponges' removal, teaser rams were used to estrus check and remained with the ewes for 96 h. The artificial insemination was made with fresh semen 10 h after estrus detection. The effect of the treatment was not significant for the estrous rates among the groups: 73%, 90%, and 86%, respectively. The main effect was observed in the pregnancy and lambing rates among the groups: 70%, 86%, 56%, and 80%, 120%, 56%, respectively. Based on these results from our study, the use of the MAP—eCG is the best choice to improve the fertility rate on ewes.

167 EVALUATION OF SUPEROVULATORY RESPONSE IN Santa Inês EWES BY ULTRASONOGRAPHY AND LAPAROSCOPY

The development and use of non-invasive techniques would reduce risks of surgery sequels on the same animal and use of the donor could be optimized. The aim of this study was to evaluate the accuracy of ultrasonography performed prior to embryo collection to estimate superovulation response in sheep. Fifteen pluriparous Santa Inês sheep, 2 to 5 year old, with an average body weight of 46.79 ± 6.00 kg and body condition score of 2.96 ± 0.32 (1 to 5 scale) were allocated into 3 groups (GI, GII, GIII) in a cross-over design. In GI, intravaginal sponges (60 mg of medroxyprogesterone acetate; Progespon®, Schering Plough Animal Health, São Paulo, Brazil) were inserted (Day 0) and maintained for 14 days, and the superovulatory (SOV) protocol started on Day 12. In GII and GIII, intravaginal sponges were inserted (Day 0) and maintained for 6 days. On Day 5, the animals were treated with 300 IU of eCG (Novormon 5000®, Schering Plough Animal Health) and 5 mg of dinoprost (Lutalyse® Pfizer Animal Health, São Paulo, Brazil) i.m. Animals in GIII received 0.025 mg of gonadorelin acetate (Gestran-Plus®, Tecnopec, São Paulo, Brazil) i.m. 12 h after sponge withdrawal. In GII and GIII, the SOV protocol started 48 h after sponge removal and a new sponge was inserted immediately after its removal. The SOV protocol in all groups consisted of 200 mg of pFSH (Folltropin-V®, Tecnopec) administered in 6 decreasing doses given every 12 h (50/50, 30/30, and 20/20 mg). At the time of the fifth dose of pFSH, 5 mg of dinoprost was administered i.m. and the sponges were removed. Animals were bred by a sexually mature ram twice a day until the end of estrus. Prior to embryo collection, an ultrasonographic evaluation (5 MHz, Aloka SSD-500, Tokyo, Japan) was performed to determine the number of CL present on both ovaries. The number of CL was further evaluated by laparoscopy, when it was possible to detect poor responders or even non-ovulating animals. Statistical analysis was performed using all tests at the 95% confidence interval by SAEG program. Results are presented as mean ± SE. The number of CL was not different among all groups. The total number (GI, GII, and GIII) of CL determined by ultrasonography (9.09 ± 5.01) was not different (P &gt; 0.05) from that observed by laparoscopy (8.87 ± 5.25). A significant correlation (r = 0.56, P &lt; 0.0005) between evaluation performed by ultrasonography and laparoscopy was observed. These results suggest that ultrasonography can be used to determine the response to the superovulatory protocol. As embryo collection in sheep is performed mainly by surgical techniques, the implementation of non-invasive techniques such as ultrasonography could avoid unnecessary surgeries on animals that did not respond to the SOV protocol, therefore preventing early culling of embryo donors. Financial support: Fopesq/Proppi and Faperj (E26/171.065/2006).

439 OVARY ULTRASOUND EVALUATION OF Santa Inês EWES SUBMITTED TO SHORT DURATION PROTOCOLS WITH PROGESTOGENS ASSOCIATED WITH LECIRELIN AND/OR ESTRADIOL BENZOATE

In order to enhance the use of biotechnology in females it is necessary to have good knowledge of the ovarian events, and thus enable the manipulation of the estrus cycle to concentrate ovulations in an established period of time. In this experiment, we evaluated the ovary response and corpus luteum (CL) parameters in a short-duration protocol associated or not to lecirelin (L)(GnRH) and/or estradiol benzoate (EB). Twenty-four cyclic Santa InÊs ewes were divided into 4 groups: Control animals (G1; n = 6) received 45 μg of d-cloprostenol (Prolise®, Tecnopec, Sao Paulo, Brazil) on Day 0. Intravaginal sponges containing medroxyprogesterone acetate (MAP60®, Tecnopec) were inserted on Day 3. Then on Day 7 sponges were removed and 400 IU of eCG and 45 μg of d-cloprostenol were administered. G2 (n = 6) animals underwent the same protocol as G1 but with administration of 1 mg of EB on Day 1; G3 (n = 6) animals underwent the same protocol as G1 but with administration of 25 μg of L (Gestran plus®, Tecnopec) 30 h after sponge withdrawal; G4 (n = 6) animals underwent the same protocol as G1 but with administration of 1 mg of EB on Day 1 and 25 μg of L 30 h after sponge withdrawal. Ewe ovaries were assessed via transrectal ultrasound (US), and CL were measured 4 days after ovulation. On the same day blood samples were taken to measure plasma progesterone levels via radioimmunoassay. Twenty-three out of 24 animals ovulated. One ewe from G4 that did not ovulate was taken out of the experiment. The average size of the largest pre-ovulatory follicle was 7.3 mm (G1), 7.0 mm (G2), 6.8 mm (G3), and 7.4 mm (G4) (P &gt; 0.05). There were 1.8, 1.2, 2.0, and 1.4 ovulated follicles per animal in G1, G2, G3, and G4, respectively. The results for estrus start and duration after sponge withdrawal were G1: 32/38.6; G2: 37/69.3; G3: 29.3/37.3; G4: 44/68 h, respectively. Ovulation time span was G1: 65.3; G2: 88; G3: 53.3; G4: 82.4 h after sponge withdrawal. There was a high correlation of ovary mensuration and progesterone level (r = 0.64; P &lt; 0.0001) on the fourth day after ovulation. Progesterone levels after ovulation were 2.37 ng mL-1 (G1); 1.5 ng mL-1 (G2); 3.22 ng mL-1 (G3) and 1.99 ng mL-1 (G4), being higher in G3 than in G2 (P &lt; 0.05). It was seen that the EB is prejudicial to the CL function. Despite the fact that there was no significant difference of progesterone levels within G1, G3, and G4, animals in G3 displayed higher levels of progesterone; hence, there is a need for further studies using a larger number of animals and fertility test. CNPq e Tecnopec.

9 EFFECT OF FLUSHING WITH PROTEIN SOURCES OF DIFFERENT DEGRADABILITY ON OVULATION NUMBER IN Santa Inês EWES

Flushing provides sheep a diet dense in nutrients before the breeding season to increase the ovulation rate and is usually accompanied by increase in body weight (BW) and body condition score (BCS). Although the effect of flushing with high-energy diets is documented, the influence of flushing with high-protein diets on ovulation rate and on plasma urea nitrogen (PUN) is not well established. The aim of this study was to evaluate the ovulation rate of Santa Ines ewes subjected to flushing with protein sources with different ruminal degradability. Forty-eight ewes were assigned to 2 treatments in a randomized block design, based on initial BCS (2.25 to 2.5; 2.75; and 3.0 to 3.25; scale 1 = thin and 5 = fat). The treatments consisted of 2 isonitrogenous (12% crude protein) and isocaloric (3.9 Mcal of metabolizable energy per day) diets, but with different proportions of rumen-degradable protein; one with soybean meal (SM, n = 24) and the other with corn gluten meal and cottonseed meal (CG + CM, n = 24), provided for 28 d before ovulation. The BW and BCS were evaluated weekly. Estrus synchronization was initiated on Day 17 after beginning the treatment diets. An intravaginal sponge with 60 mg of medroxyprogesterone acetate (MAP-60®, Tecnopec, São Paulo, Brazil) was used for 11 d, and an injection of 300 IU of eCG (Novormon®, Syntex SA, Buenos Aires, Argentina) and 50 μg of cloprostenol (Prolise®, ARSA S.R.L., Buenos Aires, Argentina) was given 9 d after beginning the protocol. Estrus was monitored twice daily. The ovulation rate was evaluated by transrectal ultrasonography and was characterized by the disappearance of dominant follicles in the ovaries. Blood samples for determination of PUN were collected on Days 7, 14, 21, and 28 after the start of flushing. Quantitative data were subjected to ANOVA and the rate of estrus detection was analyzed by chi-square test. Treatment differences were considered significant if P &lt; 0.05. The sheep consumed the equivalent to 1.60 times the metabolizable energy requirement for maintenance and 1.86 times the requirement of protein for maintenance. Estrus detection rate (95.83 v. 83.33%), BW (3.14 ± 0.55 v. 2.94 ± 0.61 kg) and BCS (0.04 ± 0.05 v. 0.10 ± 0.06) gain before ovulation, BW (54.46 ± 1.31 v. 53.36 ± 1.44 kg) and BCS (2.78 ± 0.05 v. 2.85 ± 0.05) on the 28th experimental day, PUN (17.25 ± 0.60 v. 16.23 ± 0.66 mg dL-1), ovulation rate (2.25 ± 0.16 v. 2.24 ± 0.18) and ovulatory follicle diameter (6.61 ± 0.17 v. 6.72 ± 0.19 mm) did not differ (P &gt; 0.05) between ewes feed SM or CG + CM, respectively. Flushing with protein sources with different ruminal degradability did not alter the ovulation rate or the ovulatory follicle diameter in Santa Ines ewes. Funded by FAPEMIG, CNPq, TECNOPEC.

106 PREGNANCY RATE AFTER EMBRYO TRANSFER OF IN VIVO-PRODUCED OVINE EMBRYOS CRYOPRESERVED BY SLOW FREEZING OR VITRIFICATION

Although slow freezing is the method of choice to cryopreserve in vivo-produced ovine embryos, vitrification has became an alternative procedure mostly developed for in vitro-produced bovine embryos. The aim of this work was to compare pregnancy rates after cryopreservation of in vivo-produced ovine embryos with slow freezing or open pulled straw (OPS) vitrification method. Ewes were synchronized using intravaginal sponges containing 60 mg of medroxyprogesterone acetate for 14 d. Superovulation was performed using a total dose of 176 IU of ovine FSH (Ovagen), in 6 decreasing doses (i.m.) from Day 12 to 14 of treatment (Day 0 = sponge placing). Ewes were hand mated with 2 rams of proven fertility. Embryos were recovered 6 days after estrous detection by surgical procedure, evaluated under stereomicroscope, and randomly assigned to the cryopreservation treatments. Slow freezing was performed in D-PBS supplemented with 1.78 M ethylene glycol, 0.1 M sucrose, 4 mg mL-1 of BSA, and 20% serum. Embryos were loaded into 0.25-mL plastic straws and placed into a -7°C methanol bath chamber. After seeding embryos were cooled to -35°C at a rate of 0.5°C/min and then stored in liquid nitrogen. Thawing was performed by placing the straws in a 30°C water bath for 30 sec. Vitrification was performed by using the OPS method (Vajta et al. 1998) with minor modifications. Embryos were incubated in D-PBS supplemented with 1.78 M ethylene glycol, 1.3 M DMSO for 3 min and then transferred for 25 s in vitrification solution of D-PBS with 3.56 M ethylene glycol, 2.6 M DMSO, and 0.5 M sucrose, loaded in a 1 mL drop in the OPS, and immediately submerged into and stored in liquid nitrogen. Warming was performed in D-PBS plus 0.25 M sucrose for 5 min and then into D-PBS plus 0.15 M sucrose for another 5 min. Before embryo transfer, the presence of corpus luteum (CL) was detected by laparoscopic examination. One embryo per recipient was surgically transferred in the apical extreme of the uterine horn ipsilateral to the CL. Pregnancies were determined by ultrasonography 41 days after embryo transfer. Data were analyzed using the chi-square test. We found 47.8% pregnancy rate using slow freezing (11/23) and 43.5% pregnancy rate using OPS vitrification (10/23). Statistical differences were not detected (P = 0.09). We conclude that vitrification by OPS system, with minor modifications, is a suitable procedure for in vivo-produced ovine embryo cryopreservation.

331 EFFECT OF SUCESSIVE LAPAROSCOPIC OOCYTE RECOVERY AFTER HORMONAL TREATMENT IN CROSSBRED GOATS

Laparoscopic oocyte recovery (LOR) is a valuable tool for obtaining oocytes for in vitro embryo production. When preceded by a treatment of ovarian stimulation, this technique produces an increase in the amount of oocytes recovered. However, a little information has been found to respect the effect of successive hormonal treatments on both oocyte quantity and quality. Therefore, the objectives of this study were to evaluate the ovarian response and quantitative and qualitative COC production. Five adult crossbred goats were hormonally treated with intravaginal sponges containing 60 mg of medroxyprogesterone acetate (MAP, Progespon, Syntex, Buenos Aires, Argentina) for 11 days. In the 8th day of progestagen treatment, 50 μg of prostaglandin F2α analogue (Ciosin, Coopers, São Paulo, Brazil) was administered by i.m. injection. At this time, ovarian stimulation was initiated by the administration of 120 mg pFSH (Folltropin-V, Vetrepharm, Canada) distributed in five decreasing doses (30/30, 20/20, 20 mg), at 12-h intervals. The animals were fasted for 24 h before the laparoscopic procedure, which was performed just after the sponge removal. A laparoscopic system connected to a 22-gauge needle (WTA, Watanabe, Brazil) and a vacuum pump (Biovacuum, Biocom, Brazil) providing 30 mm Hg was used. All follicles with a size larger than 2 mm present in both ovaries were counted and aspirated. The collection medium was TCM-199 supplemented with HEPES (10 mM), heparin (20 IU mL-1), and gentamicin (40 μg mL-1). The COCs were graded based on presence of cumulus cells and cytoplasm homogeneity (I to IV). The hormonal treatment and LOR procedures were repeated three times at 14-day intervals. Data were expressed in percentage or mean ± SEM. The differences were analyzed using ANOVA and Tukey’s or Fischer’s exact test when appropriate, with P &lt; 0.05. No statistical differences were found (P &gt; 0.05) for the number of follicles obtained in each LOR session: 17.0 ± 3.91, 18.75 ± 2.59, and 18.0 ± 4.73, respectively. Repeated LOR procedures also did not affect (P &gt; 0.05) the number of aspirated follicle (15.0 ± 3.92, 15.5 ± 2.33, and 16.0 ± 4.36), resulting from the three sessions, respectively. Average recovery rates were not statistically different (P &gt; 0.05), resulting in 74.7%, 62.9%, and 64.6% between sessions. With respect to the percentage of viable COCs (GI and GII) were not observed statistical differences (P &gt; 0.05), as verified the follow values at 1st to 3rd sessions: 76.79%, 84.62%, and 74.19%. In conclusion, three successive hormonal stimulation LOR procedures did not cause detrimental effects on quantitative and qualitative oocyte production, suggesting that this protocol can be used for programs of in vitro goat embryo production. This study was supported the following Brazilian agencies: FINEP, CNPq, FUNCAP, and CAPES.

447 INDUCTION OF ESTRUS IN ANGLO-NUBIAN GOATS IN THE TRANSITION SEASON WITH SHORT-TERM PROGESTAGEN PROTOCOLS WITH OR WITHOUT eCG ADMINISTRATION

Synchronized estrus can be induced by different hormonal protocols including prostaglandin, progestagens, and gonadotrophins. Equine chorionic gonadotrophin is a FSH-like hormone produced by pregnant mares that can evoke antibody formation in goats. Thus, if possible, restriction of its use should be considered. The aim of this study was to investigate the possibility to omit the administration of eCG to induce estrus in Anglo- Nubian goats during the transition season. This study was done in February of 2009 in Espírito Santo do Pinhal/SP (22°57′S latitude and 46°58′W longitude), Brazil. The local breeding season extends from March to June. A total of 10 nulliparous and 10 pluriparous Anglo-Nubian goats were equally assigned into 2 treatments with (T1) or without (T2) 200 IU eCG (Novormon 5000®, Intervet Schering Plough Animal Health, São Paulo, Brazil) i.m. administration 24 h before sponge removal. All goats received (Day 0) intravaginal sponges (60 mg of MAP; Progespon®, Intervet Schering Plough Animal Health) and 50 mg d-cloprostenol (Ciosin®, Intervet Schering Plough Animal Health) i.m. Estrous behavior and transrectal ultrasonography (5-MHz transducer; Aloka SSD 500®, Tokyo, Japan) were performed at 12-h intervals (0600 and 1800h) until 96 h after sponge removal. Statistical analysis was performed using all tests at the 95% confidence interval with a SAEG® program (Funarbe, Viçosa, Brazil). Estrous response was 100% (10/10) for T1 and 70% (7/10) for T2 (P &gt; 0.05). All animals that were detected in estrus ovulated, and none of the 3 that did not show estrus ovulated. The interval from sponge removal to estrous onset (h) was 36.0 ± 9.8 (T1) and 34.8 ± 10.5 (T2), whereas its duration (h) was 27.4 ± 14.5 (T1) and 32.6 ± 12.7 (P &gt; 0.05). The interval from sponge removal to ovulation (h) was 67.0 ± 8.0 (T1) and 73.9 ± 6.4 (T2). Also, the interval from estrous onset to ovulation (h) was 25.2 ± 8.85 (T1) and 30.9 ± 6.4 (T2). No difference (P &gt; 0.05) was observed in the number of ovulations for T1 (1.2 ± 0.4) and T2 (1.4 ± 0.5). The diameter of the ovulatory follicle (mm) was similar (P &gt; 0.05) for T1 (6.7 ± 0.6) andT2 (6.5 ± 0.6) (P &gt; 0.05). Further studies should be done to consider if the use of eCG is necessary in the transition season for Anglo-Nubian goats. Financial support: Embrapa Goats and Sheep; Capritec; CNPq; Intervet Schering Plough Animal Health; Fapemig.

413 EFFECT OF FLUSHING WITH DIFFERENT DEGRADABILITY PROTEIN SOURCE ON THE PLASMA CONCENTRATIONS OF N-UREA AND PROGESTERONE IN Santa Inês SHEEP

Protein excess in diet and or rumen degradable protein (RDP) has been associated with elevated concentrations of N-urea in blood and reduction of the concentration of progesterone (P4) in plasma. The objective of this study was to evaluate the overnutrition (flushing) effects of diets containing different rates of RDP on the plasma concentrations of N-urea and P4 in Santa InÊs ewes. Twelve ewes were assigned in a randomized block design formed by 4 body weight bands. Two isocaloric (3.10 and 2.89 Mcal/day) and isonitrogenous (13.15 and 13.46% crude protein; CP) diets with different proportions of RDP (8.43 and 6.60%) containing soybean meal and corn gluten meal, respectively, and 1 maintenance diet (1.91 Mcal/day; 11.30% CP and 6.31% of RDP) were provided for 28 days before ovulation and during the next estrous cycle. Hence, 3 treatments were compared and 4 replicates were done. On Day 17 after starting the diet treatments, intravaginal sponges containing 60 mg of medroxyprogesterone acetate (MAP-60®, Tecnopec, São Paulo, Brazil) were inserted for 11 days. Nine days after sponge insertion, 240 IU of eCG (Novormon®, Syntex S.A., Buenos Aires, Argentina) and 50 μg of cloprostenol (Prolise®, ARSA S.R.L., Buenos Aires, Argentina) were administered i.m. Blood samples for determination of N-urea were collected on Days 7, 14, 21, and 28 after starting the diets and on Days 3, 6, 9, 12, and 15 after ovulation. Blood samples collected during the postovulatory interval were also analyzed for P4. Data were analyzed by ANOVA and treatment means were compared by orthogonal contrasts, where theF-testwas significant. Contrast 1 =T1 andT2 v. T3, and contrast 2 = T1 v. T2. The N-urea and P4 concentrations in plasma were analyzed according to split plot in time scheme. The sheep dry matter (DM) and CP consumption that were recorded in the flushing diet was higher (P &lt; 0.01) than in ewes receiving the maintenance diet. The DM and CP consumption of ewes did not differ between treatments T1 and T2. The plasma N-urea concentration was not different (P &gt; 0.05) between the ewes that consumed the flushing diet and ewes that received the maintenance diet, but it was higher (P &lt; 0.01) in ewes that were fed the flushing diet containing soybean meal (25.09 mg dL-1) as compared with that of ewes fed the flushing diet containing corn gluten meal (20.81 mgdL-1). The P4 concentration in plasma did not differ (P &gt; 0.05) among animals that received the flushing diet and those that received the maintenance diet, but there was a significant difference (P = 0.05) between flushing diets containing soybean meal (7.28 ngmL-1) compared with those containing corn gluten meal (4.46 ngmL-1). In summary, the nutritional level (flushing v. maintenance) did not alter the concentrations of N-urea and P4 in plasma of ewes, but the supply of a greater protein degradability source did increase their concentrations. Funded by FAPEMIG, CNPq, and TECNOPEC.

23 INDUCTION OF ESTRUS IN CYCLIC ALPINE GOATS WITH SHORT-TERM PROGESTAGEN PROTOCOLS WITH OR WITHOUT eCG ADMINISTRATION

Depending on the season, estrus can be efficiently induced using male effect, melatonin implants, light controlled programs, prostaglandin, progestagens, and gonadotrophins. The eCG is the most used hormone employed in estrus induction protocols in goats. However, as eCG is a foreign protein, a humoral immune response (i.e. antibody formation) is displayed in goats, which tends to reduce the efficiency of eCG when successively administered. The objective of this study was to evaluate the possibility of omitting the administration of eCG to induce estrus in Alpine goats during the breeding season in herds submitted to successive hormonal estrus induction. This study was done in April (local breeding season) of 2009 in Florestal/MG, Brazil. Nulliparous (n = 10) and pluriparous (n = 10) Alpine goats were equally assigned to receive (Day 0) MAP 60 mg intravaginal sponges (Progespon®, Schering Plough Animal Health, São Paulo, Brazil) for 6 days and 50 μg D-cloprostenol (Ciosin®, Schering Plough Animal Health) i.m. with (T1) or without (T2) 200 IU of eCG (Novormon 5000®, Schering Plough Animal Health) i.m. 24 h before sponge removal. Transrectal ultrasound (5 MHz probe, Aloka SSD 500®, Tokyo, Japan) was done at 12-h intervals until 72 h after sponge removal. All goats underwent timed AI 55 h after sponge removal according to expected ovulation (Menchaca et al. 2007 Anim. Reprod. Sci. 102, 76-87). Statistical analysis was performed using all tests at the 95% confidence interval (SAEG® program, Funarbe, Viçosa, Brasil). Data are reported as percentage or mean ± SD. The following results for T1 or T2 animals (respectively) were observed: estrus response (80 and 80%), goats ovulating (80 and 100%), interval from sponge removal to estrus onset (46.0 ± 17.0 and 53.5 ± 18.1 h) and to ovulation (58.8 ± 6.2 and 66.0 ± 9.8 h), interval from estrus onset to ovulation (26.0 ± 8.5 and 26.0 ± 0.7 h), diameter of ovulatory follicles (7.1 ± 0.6 and 7.4 ± 0.9 mm), number of ovulations (1.7 ± 0.8 and 1.8 ± 0.9 h), intervals from sponge removal to insemination (56.9 ± 2.3 and 58.2 ± 1.2 h) and from estrus onset to insemination (10.9 ± 14.5 and 4.6 ± 16.1h) and pregnancy rate (20 and 40%). Eight goats were inseminated at 17 to 23 h after estrus onset, which resulted in 50% pregnancy rate, and the other 2 pregnant goats were inseminated without being detected in estrus. In Brazil, dairy goats are inseminated with frozen-thawed semen 12 to 24 h after estrus onset or 42 to 55 h after sponge removal. Thus, for Alpine goats, this study suggests that earlier AI time should be avoided. However, under heat detection, AI at 18 to 24 h can be indicated. Ovarian ultrasonography of goats submitted to estrus induction gives great information of ovulation time relative to device removal and to estrus onset, which can optimize AI efficiency. In herds with successive estrus induction this kind of information could indicate an error in the common times currently used in AI programs in goats in Brazil. Financial support: Embrapa Goats and Sheep, CNPq and FAPEMIG.

407 SUPERSTIMULATORY RESPONSE WITH DECREASING DOSES OF FOLLICLE-STIMULATING HORMONE v. A SINGLE DOSE OF FOLLICLE-STIMULATING HORMONE - EQUINE CHORIONIC GONADOTROPIN DURING BREEDING AND NONBREEDING SEASON IN SHEEP

The aim of the study was to assess possible variations in superovulatory yields due to different FSH treatments at 2 times of the year. Superovulation and embryo recovery were performed during the breeding (n = 63) andnonbreeding (n = 46) seasons in Merino ewes located at 41°S latitude. Animals were kept under the same conditions, housed outdoors in a sheltered and covered pen, and were fed a liveweight maintenance ration. All animals received 60-mg medroxyprogesterone acetate intravaginal sponges (Progespon®, Syntex, Buenos Aires, Argentina) for 14 days. Ewes were then randomly assigned to 2 different superovulatory treatments: classic (n = 74) and one shot (n = 35) in both seasons. Classic superovulatory treatment consisted of 7 decreasing doses (2 × 48 mg, 2 × 24 mg, 2 × 20 mg, and1 × 16 mg NIH-FSH-P1)ofFSH (Folltropin®-V, Bioniche, Belleville, Ontario, Canada), administered twice daily from 48 h before to 24 h after pessary removal. A dose of eCG (300 IU; Novormon®, Syntex) was administered at progestagen removal. One shot superovulatory treatment consisted of a single dose of FSH (70 mg NIH-FSH-P1) plus 300 UI of eCG injected at pessary withdrawal. Embryo donors were inseminated by laparoscopy with frozen-thawed semen (100 × 106 spz) 12 h after the onset of estrus. Surgical embryo recovery was done on Day 7 after sponge withdrawal and embryos were graded for quality according to morphology (Grade 1 = excellent or good; Grade 2 = fair; Grade 3 = poor; and Grade 4 = dead or degenerated; IETS 1998). A 2 × 2 factorial ANOVA was used to test the main effects (season and superovulatory treatment) and interactions. There were no significant differences in the proportion of responding ewes (&gt;3 corpora lutea), ovulation rate, and recovered Grades 1 to 2 embryos between the breeding and nonbreeding season (Table 1; P &gt; 0.05). However, number of recovered ova/embryos and ova/embryo recovery rate were higher during the breeding season compared with the nonbreeding season, whereas the percentage of nonfertilized ova was lower in the breeding season than in the nonbreeding season (P &lt; 0.05). Analysis of data comparing superovulatory treatments showed that the proportion of responding ewes, ovulation rate, recovered embryos, and recovered Grades 1 to 2 embryos were lower for the one shot treatment than for the classic treatment (P &lt; 0.05). Embryo recovery rate and nonfertilization rate did not differ between treatments (P &gt; 0.05). It was concluded that there was an increase in the number of total recovered ova/embryos during the breeding season compared with the nonbreeding season, although the number of recovered good-quality embryos was not affected. The use of multiple FSH injections produced a higher number of total recovered and viable embryos in Merino sheep than the one shot superovulatory treatment. Table 1.Embryo yields in ewes submitted to superovulation