Measurement of early pregnancy factor activity for monitoring the viability of the equine embryo

The viability of embryos before flushing from donor mares (n = 5) and after transfer to recipient mares (n = 7) was monitored in mare serum by detecting early pregnancy factor (EPF) using the rosette inhibition test (RIT). The EPF activity was measured in donor mares before and after natural mating at natural estrus; after ovulation on Days 2, 5 and 8; and after embryo flushing (Day 8) on Days 8, 9, 10 and 13 after ovulation. The collected embryos were transferred immediately after flushing. The EPF activity in recipient mares were measured on the day of transfer and after embryo transfer on Days 1, 2, 3 and 5. Pregnancy was confirmed on Day 12 to 14 after embryo transfer. The mean EPF activity of donor mares was increased to the pregnant level (> an RI titer score of 10) on Day 2 after ovulation. Two days after flushing the embryos, the EPF activity of donor mares had decreased to the nonpregnant level. Among the 7 recipient mares, 3 mares were diagnosed pregnant on Day 12 after embryo transfer with ultrasound. The EPF activity of the pregnant recipient mares was increased above the minimum level observed in pregnant mares on Days 2 to 3 after transfer. However, among the nonpregnant recipient mares after embryo transfer, the EPF activity of 3 mares remained at the pregnant level only 2 to 3 d and then declined to the nonpregnant level. In one recipient mare, EPF activity did not reach the pregnant level throughout the sample collection. The results of this study indicated that equine EPF can be detected in serum of pregnant mares as early as Day 2 after ovulation. From our observation, we conclude that the measurement of EPF activity is useful for monitoring the in vivo viability of equine embryos and early detection of embryonic death.     

Plasma progesterone profiles and factors affecting embryo-fetal mortality following embryo transfer in dairy cattle

The relationship between plasma progesterone (P4) levels and embryo survival, and the value of P4 profiles for the selection of cattle embryo transfer recipients is still a matter of controversy. This study reports a comparison between lactating cows and heifers (n = 407) from a single dairy herd, after transfer of either fresh or frozen-thawed good quality embryos, of their ability to sustain embryo-fetal development to term. Plasma P4 concentrations on the day of estrus (Day 0 = D0), Day 4, Day 7 and on Day 21 were measured and related to embryo survival. Plasma P4 levels on Days 0, 4 and 7 were similar in recipients later found pregnant or open. Plasma P4 levels on Day 7 were significantly higher (P < 0.01) in heifers than in cows, but they were similar in pregnant and nonpregnant heifers and in pregnant and nonpregnant cows. Pregnancy rates for fresh and frozen-thawed embryos were higher in heifers than in cows, but the differences did not reach significance. However, the overall late embryonic mortality was significantly higher (P < 0.01) and the calving rate for frozen-thawed embryos was significantly lower (P < 0.05) in cows than in heifers. As expected, plasma P4 on Day 21 was significantly higher (P < 0.001) in pregnant than in nonpregnant recipients, but there was no difference between pregnant cows and pregnant heifers. Plasma P4 levels on Day 7 of recipients presumed pregnant on Day 21 and later found pregnant or nonpregnant were similar, but plasma P4 levels on Day 21 were significantly higher (P < 0.001) in pregnant than in nonpregnant recipients. The results of this study suggest that plasma P4 levels until the day of transfer, except for the rejection of recipients with abnormal luteal function, are of limited practical use for embryo transfer recipient selection. However, in lactating cows low plasma P4 values on Day 7 might negatively affect embryo survival, while in heifers this effect is not noticeable. Lactating cows are more prone to embryo loss than heifers, especially in the case of frozen-thawed embryos; this is associated with a lower competence of the corpus luteum at Day 7.     

Accuracy of ultrasonography for pregnancy diagnosis on days 10 to 22 in heifers

Two operators independently conducted ultrasonic pregnancy examinations on nulliparous Holstein heifers on Days 10, 12, 14, 16, 18, 20 and 22, and assigned a diagnosis (pregnant or nonpregnant) and a score for degree of certainty in the diagnosis (1, 2 or 3 for low, intermediate or high, respectively). Pregnancy was retrospectively confirmed by the ultrasonographic detection of an embryo proper and embryonic heartbeat on Day 24 in 20 25 bred heifers; the five nonpregnant heifers were excluded from the analyses. Eleven nonbred heifers were included as an unequivocal source of nonpregnant heifers. Accuracy was not significantly greater than a guess (50%) before Day 18, but reached 100% on Days 20 and 22. Mean accuracy was higher (P<0.005) for nonpregnant (65 77 , 84%) than pregnant heifers (91.5 140 , 65%). For certainty score, there were main effects of day (P<0.0001), reproductive status (pregnant or nonpregnant, P<0.003), and an interaction of day and reproductive status (P<0.0001). The certainty score increased in all heifers among days and was higher (P<0.05) in pregnant than nonpregnant heifers on Days 16 to 20. For luteal area (area of corpus luteum, excluding area of fluid filled center, if present), there were significant main effects of day, reproductive status and a day by status interaction (P<0.0001 for each). Luteal area was approximately constant in pregnant heifers, but in nonpregnant heifers it was lower (P<0.05) on Days 16 to 22 than on Days 10 to 14. Uterine echotexture was scored on a scale of 1 to 3, characteristic of a diestrus, intermediate and estrus uterus, respectively. There were main effects of day and reproductive status (P<0.0001 for each) and an interaction of day and reproductive status (P<0.025). Uterine echotexture was approximately constant in pregnant heifers, but in nonpregnant heifers it was higher (P<0.05) on Days 16 to 22 than on Days 10 to 14. Pregnancy diagnosis on Days 10 to 14 was based on detection of the conceptus; however, detection of the conceptus was not accurate prior to visualization of the embryo proper (mean Day 22, range Days 20 to 24). In nonpregnant heifers, a correct diagnosis with high certainty was made when a small corpus luteum and uterine echotexture characteristic of estrus were detected. In the absence of these changes on Days 18 to 22, a diagnosis of pregnancy was made with high accuracy and intermediate or high certainty.     

Luteotrophic influence of early bovine embryos and the relationship between plasma progesterone concentrations and embryo survival

This study was carried out to evaluate the luteotrophic influence of early (before Day 7 as well as after Day 7; Day 0=estrus) bovine embryos and the relationship between plasma progesterone (P4) concentrations and embryo survival. Virgin Holstein dairy heifers (n=325) from a single herd were randomly allocated to be nonbred, bred by artificial insemination (AI) or by embryo transfer (ET). Bred heifers were either treated with 1500 IU human chorionic gonadotrophin (hCG) on Day 7 of the estrous cycle or received no hCG treatment. Plasma P4 concentrations on Days 0, 5, 7, 10, 13, 15, 17, 19 and 21 were similar in pregnant AI- and ET-bred heifers and, this was observed in both hCG-treated and untreated females. Nonbred, AI- and ET-bred nonpregnant heifers (both hCG-treated and untreated) presented similar plasma P4 concentrations. Plasma P4 concentrations of pregnant heifers significantly deviated from those of nonpregnant and nonbred heifers on Day 17. In hCG-treated heifers, plasma P4 concentrations and Day 28 pregnancy rate were significantly higher in females with an induced accessory corpus luteum (CL) than in those females without an induced accessory CL. Treatment with hCG, although inducing the formation of accessory CL and significantly increasing plasma P4 concentrations had no significant effect on Day 28 pregnancy rate. In conclusion, this study does not support the existence of any peripherally detectable luteotrophic influence from early embryos (Days 5-7). Plasma P4 was only significantly related to embryo survival on Day 17, the time of expected onset of luteolysis.     

Embryonic development in superovulated dairy cattle exposed to elevated ambient temperatures between Days 1 to 7 post insemination

Holstein heifers (n = 29) were used to determine whether thermal stress during the first 7 d of embryonic development may increase the incidence of embryonic abnormalities in dairy cattle. Heifers were acclimated to environmental chambers at 20 degrees C for 9 d and superovulated with follicle stimulating hormone-pituitary (FSH-P; 40 mg total), beginning on Days 9 to 11 of the estrous cycle. Prostaglandin F(2)alpha (Lutalyse; 50 mg total) was administered on Day 3 of FSH-P. Heifers were inseminated artificially at estrus and then maintained at either thermal neutrality (20 degrees C) or under hyperthermic conditions (daily exposure up to 16 h at 30 degrees C and 8 h at 42 degrees C) for 7 d beginning at 30 h after the onset of estrus. On Day 7 post estrus, embryos were recovered nonsurgically and evaluated morphologically for stage of development and quality. The distribution of embryos classified as normal, abnormal, retarded or as unfertilized ova, differed (P<0.001) between heat stress and thermoneutral treatments. Only 20.7% of 82 embryos recovered from stressed heifers were normal compared with 51.5% of 68 embryos from thermoneutral animals. Stressed heifers had a higher incidence of abnormal and retarded embryos with degenerate nonviable blastomeres. Responses indicated that thermal stress from 30 h after the onset of estrus to Day 7 post estrus increases the incidence of abnormal and retarded embryos in superovulated heifers.     

Effects of administering progesterone at selected intervals after insemination of synchronized heifers on pregnancy rates and resynchronization of returns to service

In 3 separate trials at 2 locations, dairy heifers (n = 396) were treated with a Controlled Internal Drug Release (CIDR) progesterone device for 9 d. On Day 7 of CIDR treatment, all heifers were injected with PGF(2alpha). Synchronized estruses were detected using a tailpaint and chalk (TPC) scoring system. An animal's tailhead was painted at device insertion, and this strip was covered with a contrasting color of chalk at device removal. Over all trials, 85.1% of the heifers were detected in estrus and were inseminated at 48 or 72 hours after CIDR removal. These synchronized and inseminated heifers were divided into the following treatment groups: 1) untreated controls, receiving no further treatment (n = 138); 2) post-insemination progesterone supplementation with a new (n = 59) or used (n = 29) CIDR device for Days 1 to 8 or 2 to 9, respectively, following insemination; or 3) resynchronization of return to service with a used CIDR device for Days 17 to 22 after insemination (n = 112). The pregnancy rate to first insemination in the control and resynchronized groups (Groups 1 and 3) was 46.4%, but decreased to 18.2% with the post-insemination progesterone supplementation. Resynchronization of returns to service (estrus detected 1 to 4 d following removal of second CIDR) occurred in 58.9% of all nonpregnant heifers in Group 3. In summary, CIDR devices used in conjunction with PGF(2alpha) effectively synchronize estrus in dairy heifers. Progesterone supplementation within 2 d of first insemination for 7 d suppressed fertility. Used CIDR devices inserted for Days 17 to 22 after first insemination resynchronized heifers not pregnant to first insemination.     

Influence of summer heat stress on pregnancy rates of lactating dairy cattle following embryo transfer or artificial insemination

Lactating Holstein cows were used to determine if pregnancy rate from embryo transfer (n = 113) differed from contemporary control cows (n = 524) that were artificially inseminated (AI). Holstein heifers (n = 55) were superovulated with FSH-P (32 mg total) and inseminated artificially during estrus and subsequently managed under shade structures. On Day 7 post estrus, embryos were recovered, and primarily excellent to good quality embryos (90.3%) were transferred to estrus-synchronized lactating cows. Cows were managed under conditions of exposure to summer heat stress. Pregnancy status was determined by milk progesterone concentrations at Day 21 and palpation per rectum at 45 to 60 d post estrus. Pregnancy rates of cows presented for AI (Day 21, 18.0%; Days 45 to 60, 13.5%) were typical for lactating cows inseminated during periods of summer heat stress in Florida. Pregnancy rates of embryo recipient cows were higher (P<0.001) than those of control cows (Day 21, 47.6%; Days 45 to 60, 29.2%). Summer heat stress had no adverse effect on heifer superovulatory response, but it increased (P<0.05) the incidence of retarded embryos (相似文献   

Plasma progesterone profiles and fertility status of anestrus Zebu cattle treated with norgestomet-estradiol-eCG regimen

Zebu cattle are notorious for poor fertility characterized by late maturity and long intercalving intervals attributed to a variety of factors, including genetic, nutritional and climatic. The aim of the present investigation, therefore, was to induce fertile estrus in acyclic pubertal heifers and postpartum anestrous Zebu cows by hormonal intervention. Pubertal Hariana and Sahiwal anestrous heifers (n=51) and postpartum cows (n=55) were either assigned a placebo (controls, N=6 for each breed and parity) or treated with 10-d norgestomet (3 mg) subcutaneous ear implants, with an initial injection of 3 mg, im norgestomet + 5 mg estradiol valerate, followed by 500 IU eCG at implant withdrawal (NOR-treated groups). Jugular venous plasma samples were obtained from a total of 28 animals (controls : 4 heifers and 4 cows; NOR-treated : 12 heifers and 8 cows) on Days 0 (implant insertion), 3, 7, 9 and Day 10 (implant withdrawal), every 12 h on Days 11 and 12, and then once daily on Days 17, 24 and 31. All the samples were assayed for progesterone. Almost all (97%) heifers and 81% cows were induced to estrus, the majority (92% heifers and 79% cows) within 120 h of implant removal. Synchrony of the induced estrus was better in cows, but interval to estrus and estrus duration were significantly longer in heifers (P<0.05). Post-treatment fertility, based on Day 28 nonretum rate, first service, and overall conception rates, was better in heifers (78.9, 60.5 and 73.7%, respectively) than cows (77.1, 48.6 and 62.9%, respectively), but the differences were significant only for the overall pregnancy rate (71.8% for heifers and 51.2% for cows; P<0.05). Low pre-treatment plasma progesterone values (<0.5ng/mL) were consistent with ovarian inactivity, confirming the true anestrus status of experimental animals. Controls failed to exhibit estrus and maintained low progesterone concentrations throughout the study. In treated animals, high progesterone values from Day 17 onwards suggested ovulatory estrus. These early luteal phase progesterone concentrations in nonpregnant (P=0.06) and nonpregnant, nonretum (P<0.05) animals were low in comparison with those of pregnant animals. Good fertility resulting from breeding according to estrus, inspite of variable intervals to estrus and estrus duration, advocates its advantage over fixed-time insemination in norgestomet-treated anestrous Zebu cattle.     

Effects of postinsemination progesterone supplementation on fertility and subsequent estrous responses of dairy heifers

The objective of Experiment I, replicated twice, was to evaluate whether fertility of estrus-synchronized dairy heifers could be improved by postinsemination progesterone supplementation. Estrous cycles were synchronized using two injections of prostaglandin (PG) F(2alpha) adiministered 11 days apart. Heifers displaying estrus were inseminated and assigned to control (n = 155) and treated (n = 159) groups. Treatment consisted of intravaginal insertion of controlled internal drug release (CIDR) devices for Days 7 to 13 (Day 0 = day of estrus). The conception rate for CIDR-treated heifers (57.9%) did not differ significantly from that of the controls (53.6%). The return-to-estrus rate and pattern of return estruses were not affected by treatment, but indicated that early embryonic mortality may have occurred in some of the heifers diagnosed nonpregnant. The objective of Experiment II was to evaluate if used CIDR devices were effective in resynchronizing returns to estrus in previously synchronized inseminated but nonpregnant and noninseminated heifers. Estrous cycles of dairy heifers of breeding age were synchronized with PGF(2alpha). Heifers displaying estrus were assigned to be inseminated (n = 117) or not inseminated (n = 35). All heifers were treated with 9-day used CIDR devices for Days 17 to 22 after synchronized estrus in order to resynchronize returns to estrus. Of the inseminated but nonpregnant heifers (n = 41), 78.1% were detected in estrus after CIDR removal (versus 94.3% of noninseminated heifers [n = 35]; P < 0.05) and 61.0% of the estruses occurred within 4 days of CIDR removal (versus 91.4% of noninseminated; P < 0.05). Estruses of synchronized inseminated nonpregnant heifers occurred over a longer period compared with those of noninseminated heifers (P < 0.025). The results indicate that response to the resynchronization protocol was altered by the outcome (early embryo death or failed fertilization) of the previous unsuccessful insemination, and support the hypothesis that delayed returns to estrus can be attributable to a pregnancy which was initiated but failed to establish itself. Such factors should be considered when evaluating responses of cattle to treatments designed to enhance fertility.     

Factors affecting pregnancy rate following nonsurgical embryo transfer in buffalo (Bubalus bubalis): a retrospective study

The objectives of this study were to determine the pregnancy rate and factors affecting it following nonsurgical embryo transfer in buffalo. Donor buffalo were superovulated with FSH, and embryos collected nonsurgically were evaluated for stage of development and quality. They were transferred nonsurgically to 91 recipients on Days 5 to 7 of the natural (n = 52) or induced (n = 39) estrus (estrus = Day 0). The overall pregnancy rate of 24/91(26.4%) was higher than in earlier reports for buffalo but was much lower than in cattle. Pregnancy rates were not affected by season (autumn vs winter), side of transfer (right vs left uterine horn), or type of estrus (spontaneous vs induced). The pregnancy rate was high 11/27(40.7%) when donors and recipients were closely synchronized, while it was compromised when recipients were in estrus at +12 h (1/7, 14.3%) and at -12 h (5/27, 18.5%). Asynchrony beyond 12 h on either side resulted into conception failure. The pregnancy rate tended to increase with the increase in CL size of recipients, while stage of embryonic development had no effect. The transfer of an 8-cell embryo with a 16-cell embryo led to the birth of heterosexual twins, indicating that the uterine milieu of Day 5 to 6 recipients may be tolerated by the out-of-phase 8-cell embryo, at least in the presence of a more mature embryo. Embryo quality had the greatest effect on pregnancy rate as it was higher (P < 0.005) after the transfer of Grade I than Grade III embryos (6/10, 60.0% vs 3/36, 13.9%). Assessment of returns to estrus indicated that among nonpregnant recipients, 17/67 (25.4%) embryos never matured sufficiently to prevent luteolysis through maternal recognition of pregnancy (MRP), while 14/67 (20.8%) embryos probably died following MRP. These results indicate that efforts to increase pregnancy rate following embryo transfer in buffalo should include prevention of luteolysis during the first week of transfer and a reduction in the incidence of embryonic mortality.     

Ovarian follicular populations during early pregnancy in heifers

Ovarian follicles >/=2mm were studied in 14 pregnant and 14 nonpregnant Holstein heifers by daily ultrasound examinations. There were significant differences among days, from Day 0 (day of ovulation) to Day 21, in the diameter of the largest follicle and the diameter of the second largest follicle in pregnant and nonpregnant heifers. There was an interaction of day and reproductive status (P < 0.001) for the diameter of the largest follicle. Significant differences among days were also observed in the numbers of follicles 2 to 3 mm, 4 to 6 mm, 7 to 10 mm, 11 to 13 mm, and >13 mm, and the total number of follicles >/=2 mm. There was a significant main effect of reproductive status for the number of follicles 11 to 13 mm. An interaction of day and reproductive status was observed for the number of follicles >13 mm, but not for any of the other diameter categories. The effect of reproductive status for number of follicles 11 to 13 mm and the interactions for the number of follicles >13 mm and the diameter of the largest follicle seemed due to the selective growth and ovulation of the follicle destined to ovulate in nonpregnant heifers. The differences in ovarian follicular populations between pregnant and nonpregnant heifers were attributed solely to the presence of a physiological mechanism for the selection of an ovulatory follicle in nonpregnant heifers. There were no significant differences among days for any follicular endpoint during Days 22 to 60 in the pregnant heifers.     

Use of short-term progestin treatment to resynchronize the second estrus following synchronized breeding in beef heifers

Two trials were conducted to evaluate the efficacy of short-term progestin administration to resynchronize the second estrus after artificial insemination in yearling beef heifers. In Trial 1 crossbred yearling heifers (n = 208) were synchronized with Syncro-Mate-B (SMB) and artificially inseminated (AI) between 48 and 54 h following implant removal. Implant removal is defined as Day 1. Following AI, the heifers were randomly assigned to 1 of 2 experimental groups. Group 1 heifers were fed melengestrol acetate (MGA) daily from Day 17 to 21 at a rate of 0.5 mg/head, while Group 2 control received no exogenous progestin during this period. Synchrony of estrus was defined as the 3-d period in which the highest number of heifers expressed behavioral estrus in each group. There was no difference (P < 0.05) in the pregnancy rate during the second estrus due to MGA supplementation. More MGA-treated heifers (P < 0.01) expressed estrus in a 3-d period than the controls. In Trial 2, yearling heifers (n = 108) were synchronized with 2 injections of PGF(2alpha) (second PGF(2alpha) injection is designated as Day 1) administered 14 d apart with AI 12 h after the onset of behavioral estrus. The heifers were then randomly assigned to 1 of the following 3 treatment groups after initial AI: 1) MGA fed at 0.5 mg/head daily from Days 17 to 21; 2) norgestomet administered in 6.0-mg implants from Days 17 to 21; 3) untreated control heifers. Blood samples were collected on Day 21 and analyzed for progesterone (P(4)). Elevated P(4) (> 1 ng/ml) on Day 21 indicated pregnancy to the first insemination. Synchrony among the 3 groups of heifers was similar (P > 0.10); however, the second estrus was less (P < 0.05) variable in the MGA and norgestomet treated heifers. During the resynchronized second estrus, conception rates were not affected by progestin treatment (MGA 40%, norgestomet 64%, and control 62%; P > 0.10). However, a proportion of heifers treated MGA 10% 4 36 and norgestomet 3% 1 36 expressed behavioral estrus during second estrus even though they were diagnosed as pregnant from first service by elevated P(4) levels on Day 21. We conclude that short-term use of progestin from Days 17 to 21 following AI causes closer synchrony of estrus; however, inseminating pregnant heifers that exhibit behavioral estrus may cause abortion.     

Inhibition of pregnancy with indomethacin in mature gilts and prepuberal gilts induced to ovulate

Twenty prepuberal (P) gilts, 56.5 +/- 1.1 kg body weight, were induced to ovulate with 1000 IU of pregnant mare's serum gonadotropin followed 72 h later by 500 IU of human chorionic gonadotropin (hCG), and bred by artificial insemination (AI) with 50 ml fresh pooled boar semen the day after hCG treatment (Day 0). Eighteen mature (M) gilts, 120.6 +/- 1.7 kg body weight, were bred by AI each day of estrus using pooled semen from the same boars (onset of estrus = Day 0). One-half of each group was fed the prostaglandin (PG) synthesis inhibitor indomethacin (IND), at 10 mg/kg body weight, or control (C) feed twice daily on Days 10 to 25. Blood samples taken by venipuncture on Days 10, 15, 20 and 25 were quantitated for progesterone (P4) and 13,14-dihydro-15-keto-PGF2 alpha (PGFM) by radioimmunoassay. Ovaries were examined on Day 26. All M-C gilts were pregnant, whereas 3 of 9 M-IND gilts (P less than 0.05) and none of the P gilts (P less than 0.01) were pregnant. Three of the 6 nonpregnant M-IND gilts displayed estrus on Day 21. The 3 remaining M-IND gilts had maintained corpora lutea (CL) on Day 26. Only corpora albicantia were present in P gilts on Day 26. Serum P4 concentrations for M-C gilts, nonpregnant M-IND gilts with maintained CL, and pregnant M-IND gilts were not different. Serum P4 for all nonpregnant gilts in which CL had regressed by Day 25 decreased to less than 5 ng/ml on Day 20.(ABSTRACT TRUNCATED AT 250 WORDS)     

Estrus synchronization in cattle using estradiol, melengestrol acetate and PGF

In Experiment 1, all cattle were fed MGA (0.5 mg/head/d) for 7 d (designated Days 0 to 6) and given PGF on Day 6. One-half were administered estradiol valerate (EV; 5 mg, im) on Day 0. At Location 1, a higher proportion (P < 0.005) of EV-treated heifers were detected in estrus and bred by AI between Days 7 and 13 than control heifers not receiving EV (27 of 33 versus 15 of 32), but the number of pregnancies (12 vs 10) was not significantly different. Eighty-three of 104 EV-treated and 89 of 106 control cows were inseminated, resulting in 50 and 45 pregnancies, respectively (not significant). At Location 2, cattle were similarly treated and exposed to bulls on Days 7 to 49. Fall pregnancy rate was higher (P < 0.015) for EV-treated than control heifers (44 of 48 vs 33 of 46), but was not significantly different for cows (22 of 26 vs 19 of 23). In Experiment 2, estradiol 17beta (E17beta; 5 mg, im) and progesterone (100 mg, im) were administered on Day 0 (instead of EV). In a third group (designated the PGF group), cattle were bred on Days 0 to 6, and PGF was administered on Day 6 to those not yet bred. For 213 cows, the percentage pregnant to a synchronized estrus was greater in the PGF group (72%) than in either the control group treated with MGA (49%; P = 0.005) or the group receiving MGA and E17beta (54%; P < 0.025). Fall pregnancy rates were 91, 89, and 96% for the 213 cows (not significant) and 89, 93, and 98% for 131 heifers (not significant) in the PGF, MGA and E17beta groups, respectively. In cattle without a functional CL, the average diameter of the largest follicle at Day 6 was 1 to 2 mm smaller in the E17beta + MGA group than in the MGA group (difference significant only in cows at Location 1). Combined for both locations, the synchronized pregnancy rate in heifers without a functional CL on Day 6 was higher (P < 0.05) in the E17beta + MGA group than in the MGA group (11 of 21, 52% versus 4 of 20, 20%). Estrogen treatment caused regression of ovarian follicles with emergence of a new follicular wave. Including estrogen in an estrus synchronization program utilizing MGA and PGF significantly increased fall pregnancy rate in heifers (at 1 location) and the synchronized pregnancy rate of heifers without a functional CL at the time of PGF treatment (combined for both locations).     

Accuracy of ultrasonography and pregnancy-associated glycoprotein test for pregnancy diagnosis in buffaloes

The aims of the present study were to evaluate and compare the accuracy of transrectal ultrasonography and pregnancy-associated glycoprotein radioimmunoassay (PAG-RIA) test for diagnosis of pregnancy in buffaloes. Two hundred and seventy-five buffalo cows and heifers were examined once for pregnancy diagnosis by transrectal ultrasonography using a 5 MHz linear-array transducer between Days 19 and 55 after mating. After ultrasound scanning, a blood sample was withdrawn from jugular vein of each animal for measuring pregnancy-associated glycoprotein using a heterologous double-antibody RIA. Based on palpation of the uterus per rectum at Days 75-90, 87 animals were designated pregnant and 188 as non-pregnant. The sensitivity of transrectal ultrasonography at Days 19-24 was 44.4%, reaching 100% from Day 31 after mating. The specificity of transrectal ultrasonography ranged between 92.5 and 100% from Days 19 to 55 after mating. The sensitivity of PAG-RIA test was 11.1% at Days 19-24 and reached 100% from Day 31 after mating. The specificity of PAG-RIA test ranged from 90 to 100% from Days 19 to 55 after mating. There were no significant differences between the sensitivity and specificity of the two tests in all examined periods. In conclusion, transrectal ultrasonography and PAG-RIA test are highly accurate tests for detecting pregnant buffaloes from Day 31 after mating onwards.     

Uterine secretion of prostaglandin F2 alpha in response to oxytocin in ewes: changes during the estrous cycle and early pregnancy.

Experiment 1 was conducted to determine when the ovine uterus develops the ability to secrete prostaglandin F2 alpha (PGF2 alpha) in response to oxytocin and how development is affected by pregnancy. Pregnant and nonpregnant ewes received an injection of oxytocin (10 IU, i.v.) on Day 10, 13, or 16 postestrus. Jugular venous blood samples were collected for 2 h after injection for quantification of 13,14-dihydro-15-keto-PGF2 alpha (PGFM). In nonpregnant ewes, concentrations of PGFM increased following oxytocin on Day 16 but not on Day 10 or 13. Concentrations of PGFM did not increase following treatment on Day 10, 13, or 16 in pregnant ewes. Therefore, the ability of oxytocin to induce uterine secretion of PGF2 alpha develops after Day 13 in nonpregnant but not in pregnant ewes. Experiment 2 was conducted to precisely define when uterine secretory responsiveness to oxytocin develops. Pregnant and nonpregnant ewes received oxytocin on Day 12, 13, 14, or 15. In nonpregnant ewes, concentrations of PGFM increased following treatment on Days 14 and 15, but not earlier. Peripheral concentrations of progesterone showed that uterine secretory responsiveness to oxytocin developed prior to the onset of luteal regression. As in experiment 1, the increase in concentrations of PGFM following administration of oxytocin was much lower in pregnant than in nonpregnant ewes; however, some pregnant ewes did respond to oxytocin with an increase in PGFM. In experiment 3, pregnant ewes received an injection of oxytocin on Day 18, 24, or 30 postmating. Concentrations of PGFM increased following oxytocin on Days 18 and 24. The conceptus appears to delay and attenuate the development of uterine secretory responsiveness to oxytocin.     

Synchronization of estrus in early diestral dairy heifers with Prostaglandin F(2)alpha and estradiol benzoate

Following observation of estrus, 134 Holstein heifers were given injections of Prostaglandin F(2)alpha (PGF(2)alpha) between Days 5 and 10 of their cycle (estrus = Day 0). They were then randomly assigned to either a group receiving 400 mug of estradiol benzoate (E(2)B) 40 h or maintained as controls. Heifers observed in estrus within 120 h of PGF(2)alpha administration were inseminated (approximately 12 h after initial observation of estrus). Blood samples for progesterone determination were drawn from the coccygeal vein on Days 15 and 21 after insemination. Pregnancy was confirmed by palpation per rectum between Days 5.0 and 60 post insemination. When control and treated heifers were compared it was found that a higher percentage of heifers treated with E(2)B exhibited estrus after PGF(2)alpha, but there had been no effect on subsequent progesterone concentrations or pregnancy rates.     

Production efficiency of Japanese black calves by transfer of bovine embryos produced in vitro

The objective of this study was to examine the production efficiency of Japanese Black beef calves after transfer of bovine embryos derived from an in vitro procedure. In vitro-produced (IVP) embryos were obtained from in vitro maturation and fertilization and in vitro development by co-culture with cumulus cells until 7 or 8 days after insemination. In vivo-developed (IVD) embryos from superovulated Japanese Black heifers and cows 7 days after artificial insemination were used as a control group. Bovine embryos were transferred nonsurgically to recipient cows on Day 7 +/- 1 of the estrous cycle. Pregnancy was diagnosed by palpation per rectum at Day 60 to 70 after estrus. Pregnancy, abortion, perinatal accident and birth rates were examined according to the origin of embryos (IVP or IVD), the number of transferred embryos (single or twin) and the storage status (fresh or frozen-thawed). In Experiment 1, production efficiency by twin transfer of fresh IVP embryos was examined. Higher pregnancy rates (52 1% vs 42 9%, P < 0.05) and birth rates (47.0% vs. 33.0%, P < 0.05) were obtained by twin transfer than by single transfer of fresh IVP embryos. Thus, the twin transfer of fresh IVP embryos was effective for production of calves, although the birth rates for single and twin transfers of fresh IVD embryos were still higher (55.5% and 76.1%, P < 0.05). But the abortion and perinatal accident rates for twin transfer of fresh IVP embryos were also significantly greater than those for single and twin transfer of fresh IVD embryos (P < 0.05). In Experiment 2, production efficiency by twin transfer of frozen-thawed IVP embryos was examined. Either single or twin transfer of frozen-thawed IVP embryos resulted in a similar pregnancy rate (41.3% vs. 46.7%, P > 0.05) and birth rate (34.1% vs. 41.1%, P>0.05). Thus, in combination with frozen-thawed IVP embryos, the twin transfer did not enhance production efficiency. In conclusion, Japanese Black beef calves could effectively produce calves by twin transfer to Holstein recipients when using fresh IVP embryos, and by single transfer when using frozen-thawed IVP embryos.     

Resynchronization of previously timed-inseminated beef heifers with progestins

The objective was to determine the efficacy of a previously used CIDR or melengestrol acetate (MGA; 0.5mg/head/day) for resynchronization of estrus in beef heifers not pregnant to timed-AI (TAI). In three experiments and a field trial, heifers were reinseminated 6-12 h after first detection of estrus. Pregnancy diagnosis was done from approximately 25-43 days after either TAI or reinsemination. In Experiment 1, 79 heifers received a once-used CIDR from 13 to 20 days after TAI and 80 heifers were untreated controls. For these two groups, there were 34 and 35 heifers, respectively, not pregnant to TAI; median +/- S.E. intervals from TAI to onset of estrus were 22 +/- 0.2 days versus 20 +/- 0.6 days (P < 0.001); estrus rates were 70.6% versus 85.7% (P = 0.1); conception rates were 62.5% versus 76.7% (P < 0.3); and pregnancy rates were 44.1% versus 65.7% (P = 0.07), for CIDR and untreated (control) groups, respectively. In Experiment 2, heifers (n = 651) were TAI (Day 0) and 13 days later randomly assigned to one of seven groups (n = 93 per group) to receive a once-used CIDR (three groups; Days 13-20), MGA (three groups; Days 13-19), or no treatment (control group). Groups given a CIDR or MGA also received: no further treatment (CIDR or MGA alone); 1.5mg estradiol-17beta (E-17beta) and 50 mg progesterone (P4) in 2 mL canola oil on Day 13; or E-17beta and P4 on Day 13 and 0.5 mg E-17beta on Day 21 (24 h after CIDR removal or 48 h after the last feeding of MGA). Pregnancy rate to TAI was lowest (P < 0.05) for the group given a CIDR plus E-17beta and P4 on Day 13 and E-17beta on Day 21. Variability in return to estrus was greater (P < 0.001) in the control and MGA groups than in CIDR groups. Conception and pregnancy rates in heifers given a CIDR (65.1 and 61.4%) were higher (P<0.01) than those fed MGA (49.6 and 40.4%), but not different from controls (62.2 and 54.9%, respectively). In Experiment 3, 616 heifers received a once- or twice-used CIDR for 7 days, beginning 13+/-1 days after TAI, with or without a concurrent injection of 150 mg of P4 (2 x 2 factorial design). Pregnancy rate to TAI was 47.2%. In heifers that returned to estrus, there was no significant difference between a once- or twice-used CIDR for rates of estrus (68.8%, P < 0.3), conception (65.9%, P < 0.6) and pregnancy (45.3%, P < 0.8). Injecting progesterone at CIDR insertion increased the median interval from CIDR removal to onset of estrus (P < 0.05) and reduced rates of estrus (63.8% versus 73.8%, P<0.05), conception (60.5% versus 70.6%, P = 0.1) and pregnancy (38.6% versus 52.2%, P < 0.02). In a field trial, 983 heifers received a once-used CIDR for 7 days, beginning 13 +/- 1 days after TAI. Pregnancy rate to TAI was 55.2%. The median (and mode) of the interval from CIDR removal to estrus was 2.5 days. Estrus, conception and pregnancy rates were 78.2, 70.3 and 55.0% (overall pregnancy rate to TAI and rebreeding, 78.7%). In summary, a once- or twice-used CIDR for 7 days, starting 13 +/- 1 days after TAI resulted in the majority of nonpregnant heifers detected in estrus over a 4-day interval, with acceptable conception rates; however, injecting progesterone at CIDR insertion significantly reduced both estrus and pregnancy rates, and estradiol treatment after CIDR removal was associated with a decreased pregnancy rate to TAI. Fertility was higher in heifers resynchronized with a once-used CIDR than with MGA.     

Immunosuppressive activity associated with early pregnancy in the bovine

Immunosuppressive activity was assessed in uterine flushings (UF) and uterine vein serum and plasma from nonpregnant and early-pregnant cows, and in media from the short-term culture of Day 18 bovine embryos. The preparations were tested for their ability to inhibit [3H] thymidine (3H-TdR) incorporation into phytohemagglutinin-stimulated bovine lymphocytes. On Days 2-3 (called Day 3), Days 9-10 (called Day 10), and Days 17-19 (called Day 18) of the estrous cycle (estrus = Day 0) and pregnancy, untreated and superovulated cows were anesthesized and jugular vein and uterine vein blood was collected. The uteri were removed and flushed to obtain UF and embryos. Uterine flushings were concentrated and tested for immunosuppressive activity at 400 micrograms uterine protein/ml culture fluid. Uterine flushings from both Day 18 pregnant and Day 18 nonpregnant cows were immunosuppressive (8/8), whereas Day 10 UF were usually not immunosuppressive (7/10). Day 3 UF were usually stimulatory or only marginally suppressive (8/8). Uterine vein serum and plasma from Day 18 cows were not suppressive when compared to jugular vein serum or plasma from the same cow; neither were Day 18 uterine vein serum or plasma suppressive when compared to those same samples taken from Day 3 cows. Embryo culture media obtained from the 48-h culture of Day 18 embryos was consistently suppressive. The activity was lost after dialysis in 1000-Mr cut-off tubing, removed by charcoal, and reduced by protease digestion. These results suggest two mechanisms whereby the embryo could escape immune rejection: 1) the progesterone-induced secretion of a uterine immunosuppressive substance(s) and 2) the production by the embryo of a low molecular weight immunosuppressive substance(s).