Early pregnancy diagnosis in sheep by progesterone and pregnancy-associated glycoprotein tests

The aim of this study was to compare the accuracy of the progesterone (P4) and pregnancy associated glycoprotein (PAG) tests for determination of early pregnancy in sheep. Estrus was synchronized in 182 Awassi x Merino ewes and blood samples were collected at Days 0 (day of the insemination), 18, 22, 29, 36, and 50 after artificial insemination (AI). Plasma P4 concentrations at Days 0 and 18 were determined by double antibody radioimmunoassay, while PAG concentrations at Days 22, 29, 36 and 50 were determined by a heterologous, double-antibody radioimmunoassay (RIA) using the bovine PAG 67 kDa subunit as tracer and standard and rabbit antiserum raised against a mixture of caprine 55 and 59 kDa PAG subunits as the first antibody. The discriminatory value for diagnosis of pregnancy by the P4 and the PAG-RIA tests was > or = 1 ng/ml. Based on lambing data, the accuracy for diagnosing pregnant (sensitivity) and non-pregnant ewes (specificity) and predictivity of both tests were calculated. The sensitivity, specificity, positive and negative predictive values for P4 and PAG tests were 100, 95.4, 81.6, and 100% at Day 18 (P4) and 93.5, 100, 100 and 98.7% at Day 22 (PAG), respectively. For diagnosis of non-pregnant ewes the PAG test had significantly higher specificity than the P4 test (P < 0.05). It is concluded that ovine pregnancy can be reliably diagnosed at Day 22 after AI by using a heterologous radioimmunoassay of PAG.     

Early pregnancy diagnosis by transrectal ultrasonography in ewes

Two studies were performed to evaluate early pregnancy diagnosis by transrectal ultrasonography (TRUS), using a 7.5-MHz transducer in ewes. In the first study, the objectives were to determine the earliest date that a reliable pregnancy diagnosis could be made (percentage of ewes detected pregnant between days 15 and 20 after mating). In the second study, the objective was to confirm the findings of the first study using a randomized controlled trial. In both studies, the ewes were restrained in dorsal recumbency, using a special chute that maintained the pelvis at an angle of 30–35° lower than the head. In the first study, 30 Suffolk ewes were synchronized and maintained with 2 rams for 5 days. Each ewe was subjected to the first TRUS on day 15 after mating and daily thereafter until day 20 (estrus = day 0). Pregnancy was defined as the presence of an embryo or extra-embryonic membranes. The percentage of ewes detected pregnant at days 15, 16, 17, 18, 19 and 20 were 0% (0/30), 26.7% (8/30), 86% (24/30), 90% (27/30), 96.7% (29/30) and 100% (30/30), respectively (P < 0.001). In the second study, 390 TRUS examinations (TRUS-1) were performed on ewes from 10 to 50 days after mating in a breeding program (group mating, hand mating, cervical and intrauterine AI; breeding group; n = 270) or with vasectomized rams (vasectomized group; n = 120). The breeding date and the status of breeding were unknown to the operator. Thirty of these ewes were mated with vasectomized rams and used repetitively four times as the non-pregnant control group. All females had a subsequent TRUS (TRUS-2) between 7 and 30 days after the TRUS-1 examination. The second TRUS was used as the standard test against which the performance of the TRUS-1 was compared. The percentage of ewes correctly diagnosed at day 15 or less, days 16, 17, 18, and 19 in the breeding group were 0% (0/29), 31.3% (5/16), 40% (8/20), 70% (7/10), and 100% (14/14), respectively (P < 0.001). All the diagnoses of ewes more than 20 days following mating in the breeding group, were correctly predicted (n = 181), as well as all ewes from the vasectomized group (n = 120). It could thus be concluded that the earliest pregnancy diagnosis using a 7.5-MHz transducer by transrectal route based on the presence of positive signs of pregnancy is at day 16 and the maximum sensitivity and negative predictive value was reached at day 20 following breeding.     

Detection of pregnancy in sheep by radioimmunoassay of sera for pregnancy-specific protein B

A radioimmunoassay (RIA) for bovine pregnancy-specific protein B (bPSPB) has been shown to be a reliable test for pregnancy in cows. Pregnant ewes have a blood antigen that cross-reacts in this RIA. Two studies were conducted to determine the accuracy of detection of pregnancy in sheep using the bPSPB RIA. In Study 1, 33 ewe lambs were bred over a 70-d period in late fall. At 26, 56, and 83 d after the end of the breeding period, blood samples were collected for assay in the bPSPB RIA, and the Pregmatic 3 ultrasonic device was used to detect pregnancy. Pregmatic 3 detected pregnancy in 14, 27 and 28 ewes and nonpregnancy in 19, 6 and 3 ewes at Days 26, 56 and 83 past the breeding period, respectively. The bPSPB assay detected pregnancy in 32, 31 and 30 ewes and nonpregnancy in 1, 2 and 2 ewes at Days 26, 56 and 83 past breeding, respectively, Thirty ewes lambed and three did not. In Study 2, 180 multiparous ewes were bred over a 60-d period in summer. At 35 d after the end of the breeding period, blood samples were collected for assay in the RIA, and a real-time ultrasonic scan was done to detect pregnancy. Real-time ultrasonic testing detected pregnancy in 163 ewes and nonpregnancy in 17 ewes; whereas, the RIA detected pregnancy in 161 ewes and nonpregnancy in 19 ewes. One hundred fifty-nine ewes lambed and 21 did not. The bPSPB RIA detected pregnancy earlier and more accurately than the Pregmatic 3 ultrasonic device and was equally as accurate as the real-time scanning instrument. These studies demonstrate an accurate serological test for a pregnancy-specific antigen in sheep.     

Plasma urea nitrogen in relation to pregnancy rate in dairy sheep

The aim of this field study was to investigate the relationship of plasma urea nitrogen (PUN) with the pregnancy rate in lactating Awassi × Merino ewes. One hundred and eighty-five Awassi × Merino ewes were used in the present study. Ewes were fed a diet containing 17.4% crude protein and were milked twice a day by the milking machine. The ewes were synchronized for estrus by insertion of intravaginal sponges containing 30 mg flurogestone acetate for 14 days. At the time of sponge removal each ewe was administered eCG (600 IU). All ewes were inseminated twice with fresh semen into the external os of the cervix at 48 and 56 h after sponge removal. The day of insemination was considered as Day 0 for calculating the gestational period. Blood samples were collected from each ewe at Days 0, 18 for measurement of PUN concentrations and at Day 22 after AI for measurement of pregnancy-associated glycoprotein (PAG) by radioimmunoassay (RIA). Thirty-eight ewes (20.5%) were confirmed pregnant by PAG-RIA test at Day 22 and by ultrasonography at Day 80. The mean (±S.D.) concentration of PUN in all ewes at Day 0 was 12.7±4.6 mmol/L. There were non-significant differences in the level of PUN between pregnant and non-pregnant ewes at Days 0 (12.2±4.2 mmol/L vs. 12.8±4.7 mmol/L, respectively) and 18 (9.6±2.9 mmol/L vs. 10.4±4.0 mmol/L, respectively) after AI. Mean PUN concentrations decreased significantly from Day 0 to Day 18 after AI in both pregnant and non-pregnant ewes. By using logistic regression analysis, there was no effect of PUN concentrations on the probability of pregnancy occurrence in the studied ewes (odds ratio: 0.97; 95% confidence interval: 0.9-1.05; P=0.45). In conclusion, there was no evidence of a relationship between PUN concentration and pregnancy rate for lactating Awassi × Merino ewes in the present study because of low pregnancy rate observed.     

Diagnosis of pregnancy in the ewe at mid-gestation

Detection of the pregnancy-specific antigen, chorionic somatomammotrophin in serum was applied to the diagnosis of pregnancy in a commercial-type flock of 286 sheep 70 days after joining with rams. At the time of testing the ewes were between Days 47 and 70 of pregnancy. Based upon lambing results, the positive diagnoses were 97% correct. However, the accuracy in diagnosing non-pregnancy, which rose from 85% when all the ewes (Day 47–70) were considered to 99% after Day 55, suggest that some pregnant ewes whose stage of pregnancy was earlier than Day 55 were not being detected and were wrongly designated non-pregnant. This pregnancy test can be successfully used from Day 55, although a correct prediction of pregnancy was made in 40 ewes between Days 47 and 54.     

Improved diagnostic accuracy by repetitive ultrasonic pregnancy testing in sheep

Operator effects and instrument accuracy in using the Scanopreg ultrasonic pregnancy detector in sheep bred at synchronized estrus were studied in three experiments. In the first study, four operators tested the same 101 ewes at 60 and 80 days after breeding. The only significant difference among the four operators was that one operator consistently underestimated pregnancy. Operators did not differ in their diagnoses between days 60 and 80. In the second study, there were no differences between two operators who tested 239 ewes 90 days after breeding. In the third study, one operator tested 318 ewes 60, 70 and 90 days after breeding. The accuracy of diagnosis of pregnancy was at least 90% on each day tested; the corresponding diagnoses of nonpregnancy were 52, 76 and 79% correct. Some ewes that were initially diagnosed as nonpregnant were correctly recognized as pregnant when tested later than day 60. Most of the missed pregnancies were in ewes carrying a single lamb. A second Scanopreg test on day 90 of ewes not diagnosed pregnant on day 60 or 70 identified additional ewes as pregnant. Paired tests (days 70 and 90) recognized 99% of the ewes that eventually lambed.     

Evaluation of false transrectal ultrasonographic pregnancy diagnoses in sheep by measuring the plasma level of pregnancy-associated glycoproteins

The present study was undertaken to investigate to what extent pregnancy diagnoses made by transrectal ultrasonography could be confirmed by measurements of plasma concentration of ovine pregnancy-associated glycoproteins (ovPAG). A total of 424 Awassi x Merino ewes were synchronized for estrus and examined by transrectal ultrasonography. In Experiment 1, the ewes (n = 156) were repeatedly scanned in a standing position on d 29, 36 and 50 of gestation. Similarly, the ewes (n = 268) in Experiment 2 were scanned on d 24, 29 and 34 of gestation, but these ewes were fasted for 12 h prior to the examination and the abdominal wall of each animal was lifted up by the hands of the assistant during the scanning. Blood samples were withdrawn after each transrectal ultrasonographic examination in both experiments. Ovine PAG concentrations were measured in plasma by a heterologous radioimmunoassay and the cut-off value for pregnancy was > or = 1 ng.mL-1. Based on the lambing performance, in Experiment 1, altogether 47 false negative and 38 false positive diagnoses were made by transrectal ultrasonography in 24 and 33 ewes, respectively between d 29 and 50 of gestation. In Experiment 2, altogether 8 false negative and 13 false positive diagnoses both were made in 7 ewes between d 24 and 34 of gestation. In both experiments, all ewes with false negative diagnoses had ovPAG concentrations higher than the threshold level for pregnancy diagnosis and all ewes with false positive diagnoses had ovPAG concentrations lower than the threshold of pregnancy. Furthermore, by the PAG-RIA test all lambed or aborted ewes (n = 63) were correctly diagnosed as pregnant and with three exceptions, all non-lambed ewes (n = 361) were correctly diagnosed as non-pregnant during the examined periods of both experiments.     

Overfeeding during early pregnancy reduces peripheral progesterone concentration and pregnancy rate in sheep

The 330 Merino ewes used in the study were placed with rams at a synchronized oestrus and, on Days 2-14 after mating, the ewes were placed in a feed lot and fed daily a low, medium or high ration (25%, 100% or 200% of maintenance respectively). Progesterone supplement was given to some ewes on Days 8-14 after mating by using a device containing 340 mg progesterone. Blood samples were taken from all ewes on Day 12 for measurement of plasma progesterone concentrations. On Day 14 after mating all ewes were returned to pasture. Pregnancy rate was determined by returns to oestrus and was later confirmed using ultrasound. There was a decline in the peripheral progesterone concentrations with increasing ration. The pregnancy rate in ewes fed a high ration was significantly reduced when compared with those of ewes fed a medium or low ration (48% vs 68 and 67% respectively; P less than 0.05). In ewes fed the high ration exogenous progesterone increased the pregnancy rate from 48 to 76% (P less than 0.01). Progesterone treatment did not influence pregnancy rates in ewes fed medium or low rations. The number of fetuses per ewe pregnant was not influenced by level of nutrition or progesterone treatment.     

Accuracy of transrectal ultrasonography for determination of pregnancy in sheep: effect of fasting and handling of the animals

The present study was performed to investigate the effect of previous fasting and lifting of the abdomen of the ewes during transrectal ultrasonographic scanning on the results of early pregnancy diagnosis. Ewes of four flocks (A, B, C and D; all Awassi x Merino ewes, n = 1247 ) aged 0.7-10 years were used in this study. These ewes were estrus synchronized and artificially inseminated. From 2 weeks later onwards, fertile rams were kept with the ewes of flocks A, B and C ( n=949 ) for natural breeding, while ewes of flock D ( n=298 ) were re-inseminated 17 days later. Transrectal ultrasonography (5 MHz) was carried out in ewes of flocks A, B and C on four separate occasions but only once in ewes of flock D. For final analysis, animals were divided over two groups: ewes of Group 1 ( n=949 scans) were scanned in a standing position within the milking parlor. Animals of Group 2 ( n=764 scans) were scanned by the same operator and with the same scanning technique, but these ewes were fasted for 12h prior to scanning and the abdominal wall was lifted, just in front of the udder during scanning. The sensitivity of the test for diagnosing pregnancy at Days 18-24, 25-30, 31-40 and 41-50 was 21.8, 32.3, 63.3 and 50% in Group 1, and 46, 92.5, 92.3 and 96.8% in Group 2, respectively. Only within Group 1, the sensitivity of the test was higher in young ewes (0.7-2 years) than in older ones (>2-10 years). Significant differences were observed at scan periods Days 18-24 and Days 41-50 of gestation. It is concluded that, fasting prior to scanning and lifting the abdomen during scanning significantly improve the accuracy of transrectal ultrasonographic pregnancy diagnosis in Awassi x Merino ewes.     

Evaluation of pituitary responsiveness to LHRH as a pregnancy test in ewes

Pregnant and nonpregnant ewes were injected with luteinizing hormone-releasing hormone (LHRH). Pituitary responsiveness, based on serum luteinizing hormone (LH), and follicle stimulating hormone (FSH) concentration, 2 hr after injection was then determined for each ewe, by radioimmunoassay (RIA) and correlated with the physiological reproductive state of each ewe. The serum LH release in pregnant ewes was significantly lower than that in nonpregnant ewes. Serum LH concentrations of pregnant ewes were further categorized according to whether the ewes were multiple (ML) or single lambing (SL). The responses by ML ewes were lower for LH than the SL responses. Follicle stimulating hormone responses were not different between pregnant or nonpregnant groups. Luteinizing hormone responses between pregnant ewes which were grouped according to 3 stages of pregnancy (1 to 5, 5 to 10 and 10 to 15 weeks pregnant) were not different from each other. Pregnancy diagnoses were made based on a fixed cut-off value, to which the LH response of each ewe to 5 mug LHRH was compared. Ewes whose response fell below this cut-off were diagnosed as pregnant. Accuracy of the diagnoses were determined by known lambing data. Diagnostic accuracy ranged from a low of 60% for nonpregnant, to a high of 95% for ML ewes. Accuracy for SL ewes (64%) was lower than for the overall pregnant group (79%), as well as that for ML ewes. Doses of LHRH, higher than 5 mug per ewe, generally produced LH release in pregnant ewes which was not significantly suppressed relative to responses of nonpregnant ewes. These results lead to the conclusion that gonadotropin response to exogenous LHRH injection is not an effective tool for pregnancy diagnosis.     

Effect of eCG on the pregnancy rate of ewes transcervically inseminated with frozen-thawed semen outside the breeding season

An experiment was conducted to determine whether factors affecting pregnancy rate out-of-season are associated more with transcervical artificial insemination (T-AI) procedures or with the reproductive state of the ewe. Twenty Finncross ewes were treated with progesterone sponges, and at sponge removal (0 h) 10 ewes were treated with eCG. Blood samples were collected for LH and progesterone analyses, and follicular development was monitored using ultrasonography. Ewes were inseminated from 48 to 52 h with 200 million motile frozen-thawed spermatozoa. The incidence of estrus, LH surges and ovulation was greater (P < 0.01) and intervals to these responses were shorter (P < 0.01) in the eCG-treated ewes. The number of follicles > 5 mm was higher (P < 0.05) in eCG-treated than control ewes. Progesterone concentrations increased and remained elevated through Day 19 in 7 eCG-treated and in 1 control ewe, and these ewes were pregnant based upon ultrasonographic examination. The results demonstrate that the T-AI technique using frozen-thawed semen produces a relatively high (70%) pregnancy rate out-of-season. The pregnancy rate was found to reflect primarily the reproductive condition of the ewe.     

Resuspending ram spermatozoa in seminal plasma after cryopreservation does not improve pregnancy rate in cervically inseminated ewes

The role of seminal plasma (SP) components on the maintenance of motility, viability and fertilising ability of frozen-thawed spermatozoa is of considerable interest. However, differences observed in constituents of SP among males could explain differences in fertility obtained in vivo. Two experiments were designed to examine the effects of seminal plasma on fertility from cervically inseminated frozen-thawed semen. The objective of Experiment 1 was to investigate if source or type of SP influences pregnancy rate. Seminal plasma was collected from rams previously classified as having either High (HSP; n=3) or Low (LSP; n=3) fertility in vivo. Artificial SP (fructose/sodium solution with 10% BSA; ASP) was made. Frozen semen from the same 6 rams was thawed and inseminated (Control) or resuspended either in HSP, LSP or ASP (20% in semen) prior to insemination of ewes (n=284, over 2 farms). The overall pregnancy rate was 28.1%. Treatments (Control, ASP, HSP and LSP) were not significantly different (P>0.3). There was no difference between HSP and LSP (P>0.5), and no effect of using ASP compared to ram SP (P>0.7), on pregnancy rate. As there was no effect of SP on pregnancy rate a repeat experiment (Experiment 2) was designed to test the effect of washing and selecting motile sperm prior to resuspending in phosphate-buffered saline (PBS) containing SP on pregnancy rate. Frozen-thawed semen from each of 2 rams was centrifuged through a density gradient, pellets were centrifuged through a wash medium and the sperm concentration/ram was counted. Sperm cells were resuspended in: (1) control PBS, (2) PBS containing 30% HSP or (3) PBS containing 30% LSP to give 100 x 10(6) motile sperm in 0.25 mL. Control straws were thawed and inseminated directly. Ewes (n=223 over 2 farms) were inseminated 57 h post-sponge withdrawal and those not returning to oestrus were slaughtered 29-50 days post-insemination for pregnancy determination. In Experiment 2, the pregnancy rate for Control, PBS, HSP and LSP were 15.4%, 2.3%, 0% and 0%, respectively, for Farm 1 (P>0.05) and 17.8%, 11.0%, 3.9% and 12.4%, respectively, for Farm 2. Under the conditions of the current study, addition of SP from different donors of either High or Low fertility status to frozen-thawed ram semen post-thawing did not improve pregnancy rate in ewes. ASP had no effect on pregnancy rate in ewes when added to frozen-thawed semen. Washing and selection of motile sperm prior to resuspension in PBS with or without SP (30%) before insemination had a negative effect on pregnancy rate in cervically inseminated ewes. Hence, the addition of seminal plasma or some of its constituents to semen does not appear to improve pregnancy rate in cervically inseminated ewes.     

Transcervical artificial insemination of Australian Merino ewes with frozen-thawed semen

We compared conventional methods for laparoscopic and cervical artificial insemination (AI) to a transcervical AI procedure (Guelph System for Transcervical AI; GST-AI) for use with frozen semen in Merino ewes. The GST-AI procedure was performed by an experienced operator in Experiment 1 (771 ewes) and by 2 inexperienced operators in Experiment 2 (555 ewes). In Experiment 1, intrauterine insemination by GST-AI was achieved in 76% of the ewes. The pregnancy rate at Day 70 for ewes inseminated by laparoscopy (48%, 120 251 ) was higher (P<0.01) than for ewes inseminated by either intrauterine GST-AI (32%, 64 201 ) or cervical AI (9%, 24 256 ). The overall (intrauterine and intracervical) pregnancy rate for GST-AI was 26% (68 264 ) and was unaffected by depth of insemination within the cervix. Pregnancy rates were unaffected by ram or day of insemination. In Experiment 2, the operators achieved intrauterine inseminations by GST-AI in 43% (78 182 ) of the ewes, with a significant operator effect (P<0.01) on depth of cervical penetration. The pregnancy rate to intrauterine GST-AI (40%, 31 78 ) did not differ from that to laparoscopic insemination. The total pregnancy rate for GST-AI in Experiment 2 (19%, 34 182 ) was lower (P<0.05) than that for laparoscopic AI (39%, 72 187 ) but superior (P<0.05) to that for cervical AI (1%, 1 186 ). The GST-AI pregnancy rates were affected by depth of AI (P<0.01) and by operator (P<0.05). It is concluded that GST-AI is superior to cervical AI, and may have application in Merinos if cervical penetration rates can be improved.     

Lack of effect of transrectal ultrasonography with restraint on lambing rate and prolificacy in ewes

The objective was to determine if transrectal ultrasonography for determination of pregnancy in restrained ewes increases embryonic/fetal death or loss of pregnancy. Ten flocks (N=873 ewes) bred in either the estrous or anestrous season were randomized, into control (C) or examined (E) groups within flock. Examined ewes were placed in a tilting squeeze chute and scanned by one of three operators for pregnancy by transrectal ultrasonography once between Days 25 and 100 post-breeding. Control ewes were not subjected to handling in the squeeze chute or pregnancy diagnosis. Two operators counted embryos in E ewes in six flocks. There were no differences between E and C ewes in percent ewes lambing or lambing rate on a per flock basis. Prolificacy per flock was greater in E ewes (P=0.05; 1.53 versus 1.60, C and E, respectively) than in C ewes. Eighty-five percent of exposed ewes lambed in the estrous season, while only 62% of ewes lambed in the anestrous season (P=0.05). Overall prolificacy did not differ with season (1.60 versus 1.53), and there was no season by treatment interaction for any variable tested. Losses averaged 0.02 per E ewe exposed in the estrous season and 0.20 in the anestrous season (P<0.05). Examined ewes in this study had similar pregnancy and lambing rates to C ewes in the same flocks and prolificacy was slightly greater. Based on these data, the combination of transrectal ultrasonography with restraint is safe for pregnancy diagnosis in ewes.     

Factors affecting pregnancy rates following laparoscopic insemination of 28,447 Merino ewes under commercial conditions: a survey

The results of laparoscopic insemination of 28,447 Australian Merino ewes with semen from 468 rams were used to study factors influencing pregnancy. The overall pregnancy rate was 71.7% (20,423/28,447). Pregnancy rates varied with type of progestagen implant, type and dosage of PMSG, fresh or frozen semen, wool type and number of ewes inseminated per hour. The pregnancy rate (64.6%) obtained with Medroxy-progesterone acetate (MAP) sponges, was significantly (P < 0.01) lower than with Fluorogestone acetate 30 mg (FGA 30; 74.7%) sponges, Fluorogestone acetate 40 mg (FGA 40; 72.1%) sponges, and Controlled Internal Drug Release (CIDR-G; 71.7%) implants. A PMSG dose of 200 IU resulted in significantly (P < 0.05) lower pregnancy rates (62.4%) compared with 250 IU (72.9%), 300 IU (79.1%) and > or = 375 IU (69.4%). The mean pregnancy rate for ewes administered Folligon PMSG was 71.9%, which was significantly higher (P < 0.001) than that of ewes treated with Pregnecol PMSG (65.8%). The use of Pregnecol PMSG and MAP sponges was associated, and thus their conditional effects could not be calculated. Ewes inseminated with fresh semen were significantly (P < 0.001) more likely to become pregnant (82.2%) than those inseminated with semen frozen in pellets (69.5%) or straws (71.6%). Ewes inseminated during the months of March, April or May (fall, 71.5%) were just as likely to become pregnant as those ewes inseminated in November, December, January or February (69.6%). Significantly (P < 0.05) fewer strong wool ewes become pregnant to laparoscopic AI, (67.6%) than fine (71.7%), fine medium (73%) or medium wool ewes. Significantly (P < 0.0001) more pregnancies (77.6%) were achieved when more than 55 ewes were inseminated per hour compared with fewer than 35 ewes per hour (63.4%).     

Evaluation of the beta-Neocept test for early pregnancy.

We tested the validity of the beta-Neocept test for early pregnancy against that of the plasma human chorionic gonadotropin beta-subunit radioimmunoassay (beta-HCG RIA). The beta-Neocept test had a sensitivity of 88%, a specificity of 93%, a positive predictive value of 95%, a negative predictive value of 84% and an accuracy of 90%. In view of these performance characteristics, its low cost and its ease of use, the beta-Neocept test could be used as the initial pregnancy test when there is a high probability of pregnancy, as there was in this study population, which consisted of 111 women attending endocrine infertility clinics. The more expensive beta-HCG RIA could be reserved for special indications and for patients in whom the results of the urinary hemagglutination inhibition tests are inconsistent with the clinical signs and symptoms.     

Accuracy of evaluation of ovarian structures by transrectal ultrasonography in ewes

The objective of this study was to assess the accuracy of estimating the size and number of corpora lutea (CL) and ovarian follicles by ultrasonography (US) of the ewe in standing position. US observations were compared with those made at subsequent postmortem examination of the ovaries. Corriedale ewes (n=50) of unknown reproductive history and at random stages of the oestrus cycle were used for the study. Transrectal US was performed using a 7.5 MHz transducer with the ewe in standing position. The ewes were slaughtered 12 h after the US examination and the ovaries collected, dissected and the number and size of the CL and follicles evaluated. CL were classified as functional or non-functional on the basis of their colour and follicles were classified by size (2, 3, 4 and > or =5 mm). Accuracy of US was assessed by calculating its positive predictive value and sensitivity. The data were evaluated by Pearson correlation and linear regression analysis. The predictive value and sensitivity of US was 100% for the presence and 96% for the absence of CL. In four out of five ewes with double ovulations, the presence of both CL was correctly diagnosed by US. For functional CL, the sensitivity of US was 100%. The regression coefficient for CL diameter was significant (P< or =0.001; r2=0.4; n=35). The size of 85 of 117 follicles was accurately determined. The correlation between numbers of follicles counted by US and postmortem increased with size of follicle from r=0.44 (P< or =0.01) for 2 mm to r=0.85 (P< or =0.001) for >or =5 mm diameter follicles. The regression between the diameter of follicles determined by the two methods was significant (P< or =0.001; r2=0.8; n=117). The predictive value of US for the number of follicles was high (98-100%) for all follicle sizes except for 3 mm diameter follicles (predictive value 71%). Similarly, the sensitivity was high for all sizes of follicles (90-95%) except for those of 2 mm diameter (62%). It was concluded that ultrasound scanning provides a highly accurate method for determining the number of CL and follicles > or =4 mm diameter but that its predictive value and sensitivity are lower for smaller diameter follicles. The regression equation for diameter measured by US on that evaluated postmortem explained more of the variation for follicles than for CL.     

Pregnancy diagnosis and assessment of fetal numbers in the ewe in a commercial setting

Jugular plasma progesterone concentrations were used to accurately predict open ewes (96 +/- 3%) in early pregnancy, but they less accurately predicted subsequent lambing especially during the late breeding season and most of the seasonal anestrus. Progesterone values clearly indicated that 500 I.U. of P.M.S.G. elevated ovulation rate in synchronized ewes, but did not clearly indicate fetal numbers. During late pregnancy (88-108 days), abdominal palpation, doplar ultrasound and serum progesterone analysis were equally efficacious in predicting lambing (86 +/- 9.8%, 90 +/- 9.0% and 87 +/- 4.1%, respectively), but a high percentage of ewes lambed that were diagnosed as nonpregnant (30 +/- 15.0%, 48 +/- 17.3% and 25 +/- 8.4%, respectively). Accuracy of the serum progesterone test improved the later the test was performed, although considerable individual overlap existed. Progesterone values for ewes bearing 1, 2, or greater than 2 fetuses at 94 to 95 days of gestation differed (5.5 +/- 0.3, 8.0 +/- 0.4 and 12.4 +/- 2.1, respectively (P < 0.05), whereas at 103 to 108 days values for ewes carrying two or more fetuses did not differ.     

Effects of active immunization against a synthetic peptide sequence of the inhibin alpha-subunit on plasma gonadotrophin concentrations, ovulation rate and lambing rate in ewes.

Thirty adult Mule (Blue-faced Leicester x Swaledale) ewes were actively immunized against a synthetically produced peptide corresponding to the N-terminus of the alpha-subunit of bovine inhibin conjugated to tuberculin purified protein derivative (PPD). Primary immunization in the late anoestrous period was followed by two booster injections at 5 week intervals. Control groups were either not immunized (n = 15) or received PPD only (n = 15). Ten days after the second booster, oestrus was synchronized using progestagen sponges and ovulation rate was assessed by laparoscopy on days 9-10 of the cycle. Blood samples were taken at the time of each immunization and immediately before laparoscopy. Ewes were mated with fertile rams in mid-November and the resulting conception, pregnancy and lambing rates monitored. All inhibin-immunized ewes generated antibodies that bound 125I-labelled native bovine inhibin (M(r) 32,000), and their plasma follicle-stimulating hormone (FSH) concentrations after the second booster were significantly higher than the preimmunization values (30%; P less than 0.001) and the corresponding value in the controls (25%; P less than 0.025). Inhibin immunization was associated with a 90% increase in ovulation rate (P less than 0.005) and had no adverse effect on conception rate (100%), pregnancy rate (100%) or length of gestation (146 days). However, only a 37% increase (P less than 0.05) in lambing rate was recorded for inhibin-immunized ewes, indicating a higher incidence of wastage of ova, or embryos, or both, in these ewes.(ABSTRACT TRUNCATED AT 250 WORDS)     

Differences between Belclare and Suffolk ewes in fertilization rate, embryo quality and accessory sperm number after cervical or laparoscopic artificial insemination

Ewe breed has been shown to have a major effect on pregnancy rates following cervical AI using frozen-thawed semen. The main objective of this study was to examine the differences between purebred Belclare and Suffolk ewes (multiparous) in fertilization rate, number of accessory sperm and stage of embryo development on day 6 after cervical or laparoscopic AI with frozen-thawed semen. In experiment 1, Belclare and Suffolk ewes were synchronized for 12 days and were either cervically inseminated (year 1: n=28 and 31; year 2: n=16 and 15, respectively) or laparoscopically inseminated (year 2: n=13 and 14). In experiment 2, superovulated Belclare (n=4) and Suffolk (n=13) ewes were laparoscopically inseminated. All ewes were slaughtered 6 days after AI; oocytes/embryos were recovered, morphologically graded and stained to assess the number of cells and accessory spermatozoa. Data from both experiments were combined for statistical analysis. The proportion of ewes with fertilized oocytes was significantly higher following laparoscopic AI compared with cervical AI (54% versus 19%). More Belclare than Suffolk ewes yielded fertilized oocyte(s) after cervical AI (34% versus 10%, P<0.02) but there was no difference after laparoscopic AI (62% versus 60%). From the ewes that yielded at least one fertilized oocyte the proportion of Belclare ewes with embryos at the morula/blastocyst stage was significantly greater than for Suffolk ewes (94% versus 59%, P<0.02). A higher proportion of Belclare than Suffolk ewes had evidence of sperm reaching the site of fertilization following cervical AI (39% versus 15%, P<0.02) but there was no difference after laparoscopic AI (62% versus 64%, P>0.8). Amongst the ewes with evidence of sperm at the site of fertilization, laparoscopic AI resulted in a higher number of sperm per oocyte/embryo or per ewe than cervical AI (P<0.01). These results suggested that the difference in pregnancy rate between Suffolk and Belclare ewes following cervical AI was due to: (i) sperm traversing the cervix and uterus in a higher proportion of Belclare than Suffolk ewes, leading to a higher incidence of fertilization and (ii) the lower developmental competence of fertilized oocytes from Suffolk ewes.