Search

OBJECTIVE To assess the effect of transrectal palpation (TRP) performed with the fetal membrane slip (FMS) technique for early pregnancy diagnosis on the proportion and type of associated pregnancy losses (PLs) in dairy cattle. ANIMALS 580 healthy pregnant cattle. PROCEDURES Data for artificially inseminated females with 1 or 2 viable embryos detected by transrectal ultrasonography (TRUS) at approximately 30 days of gestation were retrospectively assessed. Cattle were assigned to 1 of 2 groups on the basis of whether they did or did not undergo TRP once between 34 and 41 days of gestation (palpation and control group, respectively). At approximately 45 and 60 days of gestation, all cattle were reevaluated by TRUS; PL was categorized as type I (FMS detectable by TRP and TRUS-confirmed evidence of embryo or fetus degeneration and a functional corpus luteum) or type II (FMS undetectable by TRP and no TRUS-confirmed evidence of an embryo or fetus or of a functional corpus luteum). RESULTS Of the 580 healthy pregnant cattle, 271 underwent TRP and 309 did not. In the palpation and control groups, PL occurred in 40 (14.8%) and 47 (15.2%) cattle, respectively. Among the palpation group's PLs, 17 (43%) were type I and 23 (58%) were type II. Among the control group's PLs, 27 (57%) were type I and 20 (43%) were type II. The prevalance and type of PL did not differ between groups. CONCLUSIONS AND CLINICAL RELEVANCE TRP with the FMS technique for early pregnancy diagnosis did not increase the prevalence of PL in dairy cattle or alert the proportion of type I versus type II PL.

The objectives of the present study were to estimate the number of colonies forming units (CFU) from penile mucosa and semen, the effect of two antiseptic solutions used to flush the preputial cavity to reduce the bacterial counts from those sites, and compare them. Six clinically healthy bulls between 15 and 16 mo old declared satisfactory potential breeders were used. A prospective, randomized, and controlled cross-over design was performed, in which each bull was first sampled from the penile mucosa and semen without treatment (control group) and 24 h later, after antiseptic preputial flushing (treated group). In the treated group, the preputial area was cleaned, the preputial hair was cut, urination was stimulated, prepuce area was scrubbed twice, and the preputial cavity was flushed with either 1% of povidone-iodine solution (POI; 500 mL) or 0.05% chlorhexidine digluconate (CHG; 500 mL), maintained for 10 min. Then, the preputial cavity was emptied and flushed with 500 mL of sterile saline solution. Next, the accessory sexual glands were massaged per rectum. Finally, protrusion, erection, and ejaculation were obtained by electroejaculation, and samples from penile mucosa and semen were collected for microbiological culture. The number of CFU was determined for each sample by enumerate total aerobic bacteria using Standard Plate Surface Count cultured for 48 h. In the first replicate, half of the bulls were treated with CHG, and the other half were treated with POI. After 58.8 ± 5.3 days (x ± SD) of wash-out period, the treatments were reverted, and the same protocols were applied again. In the control group, the median number of CFUs from the penile mucosa was 750,000 (range from 60,000 to 1,800,000) and the median number of CFUs in semen was 8,000,000 (700,000-45,000,000). The CFU in semen was higher than the penile mucosa (P = 0.005). Both antiseptic solutions reduced the median number of CFUs on the penile mucosa to 915 (P = 0.002) and in semen to 1,680 (P = 0.002). The antiseptic effect on the penile mucosa was higher for CHG solution (490) than for POI solution (6,650; P = 0.05). The antiseptic effect on semen of CHG was also greater (200) than for the POI solution (31,000; P = 0.05). It can be concluded that the median number of CFU was higher in semen compared with penile mucosa, and flushing the preputial cavity either with 0.05% CHG or 1% POI maintained for 10 min reduced the number of CFUs from penile mucosa and semen. The level of antiseptic activity was higher for CHG than for POI.

The objectives of this investigation were to determine the effect of the time of copulation during the estrus period on estrus duration and luteinizing hormone (LH) response in goats. A controlled randomized study with two replicates (first = n = 12; second = n = 24), in which Boer does were divided at each replicate into three groups, was performed. Copulations at the beginning of estrus (two copulas within the first 4 h after estrus; COP-1; n = 12), copulations around the middle of estrus (two copulas around 16 h after estrus; COP-2; n = 12), and noncopulations (only mounts) throughout the estrus period (control group; CON; n = 12) were performed. Estrus duration for CON group was 41.3 ± 8.2 h; for COP-1, it was 34.0 ± 5.3 h, and for COP-2, it was 39.7 ± 6.9 h (P = 0.04). Differences were detected between COP-1 and CON groups (P = 0.01) and between COP-1 and COP-2 groups (P = 0.05) but not between CON and COP-2 groups (P = 0.56). The LH peak time for the CON group was 20.0 ± 8.0 h; for the COP-1 group, it was 13.0 ± 3.6 h, and for the COP-2 group, it was 20.5 ± 5.8 h (P = 0.04). The COP-1 group was different than the COP-2 (P = 0.02) and CON groups (P = 0.03), and no differences were detected between these last two groups (P = 0.87). It was concluded that copulation reduced estrus duration and hastened the LH peak time only when performed during the beginning of estrus.

The objective of this investigation was to evaluate the pharmacokinetic parameters of tulathromycin in plasma and semen of beef bulls after administering a single sc dose at two different sites in the neck. Four Simmental bulls with excellent temperament received a comprehensive physical exam that included breeding soundness examination. In addition, blood was collected and analyzed for CBC and chemical panel in order to rule out any subclinical liver or kidney disease. All bulls were diagnosed as healthy and satisfactory potential breeders. The mean plasma levels of tulathromycin for the two neck sites of sc administration were not different between posterior aspect of the ear where it attaches to the head (RP; regio parotidea; 77.9 ± 43.3 ng/mL; X ± SD) and to the middle of the neck (RC; regio collis lateralis; 73.7 ± 39.7 ng/mL; P = 0.84). The mean seminal plasma levels of tulathromycin after administration in the RP was 608 ± 374 ng/mL and for RC was 867 ± 599 ng/mL without differences between both sites (P = 0.29). The mean level of tulathromycin in plasma was 75.8 ± 40.2 ng/mL, which was lower than mean seminal plasma levels of 781 ± 482 ng/mL (P = 0.001). The plasma peak tulathromycin concentration (Cmax) was 160 ± 27 ng/mL at 21 ± 6 h (Tmax) post-administration. The seminal plasma Cmax was 1539 ± 44.4 ng/mL at 33.00 ± 18.00 h (Tmax) post-administration. The Cmax between plasma and seminal plasma were different (P = 0.008) without any differences in Tmax between plasma and seminal plasma (P = 0.35). The terminal half-life for plasma tulathromycin (81.4 ± 27.6 h) showed a tendency to be shorter than in seminal plasma (114.7 ± 21.7; P = 0.10). The plasma area under the curve concentration time from the first to the last sample (AUC0–last) was 15,440 ± 1717 ng/mL/h, which was significatively smaller compared with 171,071 ± 58,556 ng/mL/h for seminal plasma AUC0–last (P = 0.01). The plasma means residence time from the first to the last sample (MRT0-last) was 89.3 ± 5.1 h and it was shorter than for seminal plasma of 96.6 ± 5.0 h (P = 0.05). From the present investigation, it was concluded that tulathromycin is a suitable antibiotic based in its pharmacokinetic properties that could be used for treatment of bull genital infections when its application is indicated.

The objective of the present study was to evaluate the effect of 24 h’ fasting prior to semen collection by electroejaculation on behavioral responses, volume of rectal fecal content, bladder size, penis protrusion, erection, ejaculation stimuli, and ejaculate parameters in young Simmental bulls. Twenty-two Simmental beef bulls with an age of 13.2 ± 1.2 mo (mean ± SD) were used in a prospective randomized blinded controlled cross-over design with two pens fasted for 24 h (n = 9; FAS group), and the other three pens were non-fasted (n = 13; CON group). The bulls were maintained under confined conditions without access to pasture. One week later, the pen treatments were inverted, and semen was collected again under the same conditions and by the same team. The behavioral responses, volume of fecal rectal content, bladder size, as well as the number of stimuli required to obtain penis protrusion, erection, and ejaculation to electroejaculation were measured. The following ejaculate parameters were measured: volume, concentration, spermatozoa motility, and morphology. The behavioral response of the bulls to electroejaculation was not different between the CON group and the FAS group (3.2 ± 0.5 and 3.0 ± 0.7, respectively; P = 0.36). Bladder size was significantly reduced in the FAS group compared with the CON group (2.3 ± 0.8 vs. 2.8 ± 0.9, respectively; P = 0.02). The volume of feces in the rectum was not different between the two groups (CON was 2.3 ± 1.7 and FAS was 3.0 ± 1.8; P = 0.23). Compared with the CON group, the FAS group showed a higher proportion of penis protrusion (100% versus 81.8%, P = 0.10), erection (100% versus 81.8%; P = 0.10), and ejaculation (100% versus 90.9%; P = 0.49). The combined efficiency of penis protrusion, erection, and ejaculation (CE-PPEE) in the FAS group was superior to that of the CON group (P = 0.001). The number of stimuli necessary for penis protrusion, erection, and ejaculation for the CON group were 13.5 ± 3.7, 14.9 ± 3.7, and 20.8 ± 5.8, and they were 15.0 ± 4.2, 16.6 ± 4.2, and 20.2 ± 8.1 for the FAS group. The number of stimuli for penis protrusion (P = 0.09), erection (P = 0.08), and ejaculation (P = 0.77) were no different between the two groups. Ejaculate volume was 4.0 ± 2.6 ml and 4.1 ± 2.3 ml for the CON and FAS groups, respectively (P = 0.90). The motility was 1.4 ± 0.7 and 1.4 ± 0.8 for the CON and FAS groups, respectively (P = 0.72). The concentration of spermatozoa was 336.2 ± 273.1 million and 421.1 ± 300.6 million for the CON and FAS groups, respectively (P = 0.31). The percentage of normal spermatozoa was 50.9 ± 18.8 and 45.6 ± 14.3 for the CON and FAS groups, respectively (P = 0.16). It was concluded that fasting for 24 h prior to semen collection by electroejaculation reduced the bladder size and increased the proportion of bulls with penis protrusion, erection, and ejaculation without any difference detected in behavioral responses, volume of rectal fecal content, and ejaculate parameters.

The objectives of these two experiments were to determine the day of onset of luteolysis after exogenous PGF-2α administration during the estrous cycle and the fertility of this synchronized estrus in goats. In the first experiment, during the breeding season, 48 Nubian does were estrous synchronized, using intravaginal sponges impregnated with a progestin, and estrus was detected by vasectomized bucks. The does were divided at random into three groups of 16 does each to be treated at days 2, 3, and 4 of the estrous cycle (estrus = day 0). Then, at each day of injection, the does were again randomly divided to receive a single dose of natural prostaglandin F-2α im (PGF-2α; 5 mg/doe; treatment [TRE] group) or sterile saline solution (control [CON] group; 1 mL/doe). Finally, the following groups were originated: TRE-2, CON-2, TRE-3, CON-3, TRE-4, and CON-4. The overall estrus response after treatment with PGF-2α (TRE group, 70.8%) was higher than saline (CON group, 12.5%, P ≤ 0.001). Estrus response for TRE-2, CON-2, TRE-3, CON-3, TRE-4, and CON-4 was 25% (2 of 8), 12.5% (1 of 8), 87.5% (7 of 8), 12.5% (1 of 8), 100% (8 of 8), and 0% (0 of 8) for the same groups, respectively. Estrus response was different between day 2 and days 3 and 4 (P ≤ 0.04) and not between day 3 and day 4 (P ≥ 0.05). In the second experiment, 15 multiparous Boer does were estrous synchronized with control internal drug release (CIDR, 300 mg progesterone = P4) and PGF-2α and randomly divided to receive one single dose of PGF-2α im at days 2, 3 or 4, after synchronized estrus (n = 5 at each day). The does were detected twice a day for estrus, and blood was collected daily for P4 determination for 11 days after the synchronized estrus. Each doe in estrus was bred by hand mating to a proven male. All the does with a functional corpus/corpora luteum/lutea (CL; ≥1.0 ng/mL of P4) responded to PGF-2α with a drop in P4 levels that either lasted only 24 h for the does that did not show estrus (0.27 ± 0.10 ng/mL; n = 4) or persisted longer in all the does that showed estrus (0.22 ± 0.18 ng/mL; n = 10; P = 0.47). Estrus response for days 2, 3, and 4 was 20% (1 of 5), 80% (4 of 5), and 100% (5 of 5), respectively (P = 0.05). The conception rate was 100%, 100%, and 80% for the same days of administration, respectively (P = 0.64). It was concluded that luteolytic action of PGF-2α begins at day 3 of the estrous cycle by inducing an ovulatory and fertile estrus in goats.

The objectives of the present investigation were to evaluate the pregnancy diagnosis by detection of either the allantochorion membrane (FMS) or amniotic sac (ASP) by per rectum palpation (PRP) during late embryonic or early fetal period on pregnancy loss (PRL) at reexamination, calving rates, and abnormalities in newborn calves. A controlled randomized blind design with 800 lactating dairy pregnant cows diagnosed by transrectal ultrasonography (TRUS) between Days 35 and 57 of gestation from one dairy farm were included. The cows were randomly divided according to detection of allantochorion membrane (FMS group; n = 264), detection of amniotic sac (ASP group; n = 266), and TRUS (control [CON] group; n = 270). TRUS was considered as the criterion standard method of comparison. The entire PRP was performed by one experienced veterinarian. Then, all the cows were reexamined only by TRUS between 2 and 4 weeks later by two independent veterinarians to assess PRL. The calving rate one (number of cows calved divided by the number of cows initially pregnant) and calving rate two (number of cows calved divided by the number of cows pregnant at reexamination) for each group was calculated. All abortions and stillborns were necropsied, and calves alive were followed for 5 days. The overall initial PRL (between initial pregnant cows and reexamination) for FMS, ASP, and CON groups was 7.4% (19/258), 8.8% (23/262), and 9.2% (24/260), respectively (P = 0.75). The overall late PRL (between reexamination and calving) for FMS, ASP, and CON groups was 4.2% (9/213), 5.7% (12/209), and 4.2% (9/216), respectively (P = 0.71). The calving rate one for FMS, ASP, and TRUS groups was 79.1% (204/258), 75.2% (197/262), and 79.6% (207/260), respectively (P = 0.63). The calving rate two for the same groups was 85.4% (204/239), 82.4% (197/239), and 87.7% (207/236), respectively (P = 0.27). The number of fetuses aborted late, premature, and mature dead from FMS, ASP, and CON groups was 6, 4, and 5, respectively (P = 0.85), and no abnormalities at necropsy were detected. One stillborn male calf with atresia coli after 281 days of gestation from a cow examined by ASP at Day 51 was diagnosed. It was concluded that the use of either FMS or ASP for pregnancy diagnosis during late embryonic or early fetal period did not increase the PRL, affect calving rates, or produce calves with congenital abnormalities.

The objectives of this investigation were to determine the effect of copulation on estrus duration, LH response and ovulation in Boer goats. A controlled randomized study, with two replicates, in which does were divided at each replicate in treatment (COP; n = 12) and control (CON; n = 12) groups was performed. All the does were pluriparous and estrus synchronized with CIDR (progesterone 300 mg) maintained in the vagina for seven days, and received 50 μg of GnRH at device insertion and 5 mg of natural prostaglandin F-2α im at CIDR removal. The COP group received two copulas within the first 4 h of estrus onset, and the CON group was only permitted to be mounted. Estrus was detected twice a day during the first 24 h after pessary removal and then every 4 h by using bucks with canvas apron as teasers, led by leash for 96 h. Blood was collected during all the estrus period after each estrus detection and analyzed for LH by radioimmunoassay (RIA). In addition, at the second replicate ovulation time and number of ovulations were also monitored by transrectal ultrasonography using a linear 7.5 MHz probe beginning 24 h after estrus onset and repeated every 4 h until all the preovulatory follicles disappeared. Estrus onset was 36.7 ± 10.5 h and 35.5 ± 13.6 h for CON and COP groups, respectively (P = 0.82). Estrus duration for the same groups was 40.3 ± 9.9 h and 28.3 ± 4.7 h, respectively (P = 0.001). The LH peak time for the CON group was 17.7 ± 6.3 h, and for the COP group, it was 10.9 ± 2.6 h (P = 0.004). The LH peak magnitude for the same groups was 31.5 ± 16.2 ng/mL and 34.9 ± 20.7 ng/mL, respectively (P = 0.34). The LH peak duration was not different between groups (CON: 7.3 ± 1.6 h versus COP: 7.2 ± 2.4 h; P = 0.94). The first ovulation time for CON and COP groups was 33.7 ± 3.9 h and 29.1 ± 3.2 h (P = 0.05), and the last ovulation time for the same groups was 37.7 ± 3.9 h and 32.6 ± 2.5 h, respectively (P = 0.02). The overall time from LH peak to ovulation was 18.6 ± 4.8 h without differences between groups (CON: 16.3 ± 5.6 h versus COP: 20.6 ± 3.3 h; P = 0.15). The number of ovulations for the CON group was 2.2 ± 0.4, and for COP group, it was 2.1 ± 0.4 (P = 0.96). It was concluded that copulation reduced estrus duration and hastened LH peak and ovulation in Boer goats.

The objectives of the present study were to evaluate the effect of per rectal amniotic sac palpation (ASP) for pregnancy diagnosis during the late embryonic period on pregnancy loss, calving rates, and abnormalities in newborn calves. A controlled, randomized, blocked, blind experiment containing 680 lactating pregnant dairy cows with a viable embryo diagnosed by transrectal ultrasonography was performed. Two dairy operation sites (farm A and farm B) were selected. At each farm, the cows were randomly divided into control (CON) and ASP groups. The CON group was not subjected to pregnancy diagnosis via per rectum palpation. The ASP examinations were performed by one experienced veterinarian between Days 34 and 45 after breeding. All cows were reevaluated by transrectal ultrasonography only between 2 and 4 weeks later. Two calving rates were calculated: calving rate 1 (cows that calved from the initial number of pregnant cows) and calving rate 2 (cows that calved from cows pregnant at reexamination). In farm A, the percentages of early pregnancy loss were 11.5% (19 of 165) and 13.2% (24 of 182) for the CON and the ASP groups, respectively (P = 0.64). In farm B, the percentage of early pregnancy loss was 11.2% (19 of 170) for the CON group and 8.8% (14 of 159; P = 0.48) for the ASP group. In farm A, the percentage of late pregnancy loss was 7.6% (11 of 145) for the CON group and 5.5% (8 of 155; P = 0.39) for the ASP group. In farm B, the percentage of late pregnancy loss was 3.7% (5 of 137) for the CON group and 6.3% (8 of 127; P = 0.32) for the ASP group. In farm A, early pregnancy loss was higher than late pregnancy loss (12.4% vs. 6.3%; P = 0.01), and in farm B, the same tendency was detected (10.0% vs. 4.9%, for early and late pregnancy loss, respectively; P = 0.02). In farm A, calving rate 1 was 81.2% (134 of 165) for the CON group and 80.8% (147 of 182; P = 0.92) for the ASP group. Calving rate 2 for the same groups was 92.4% (134 of 145) and 94.8% (147 of 155), respectively (P = 0.68). In farm B, calving rate 1 was 77.7% (132 of 170) for the CON group and 74.8% (119 of 159; P = 0.55) for the ASP group. Calving rates 2 for the same groups were 87.4% (132 of 151) and 82.1% (119 of 145), respectively (P = 0.20). Two female calves with atresia coli were diagnosed only in the CON group. It was concluded that ASP during the late embryonic period for pregnancy diagnosis did not increase the pregnancy loss, affect calving rates, or produce abnormalities in calves.

The objective of the present study was to evaluate the effect of copulation on estrus duration and ovulation in goats. During the fall season, 14 multiparous Boer does were estrous synchronized with controlled internal drug release (300 mg), maintained in the vagina for 7 days, and received 50 μg of intramuscular GnRH device insertion and 5 mg of natural intramuscular PGF2α at device removal. The does were randomly divided into two equal groups: a treatment group (TRE; n = 7) and a control group (CON; n = 7). The TRE group received two copulas by fertile bucks within the first 4 hours of estrus onset, and the CON group received only mounts by the same males equipped with canvas aprons. Estrus detection was performed every 12 hours after controlled internal drug release removal within the first 24 hours and then every 4 hours for 5 days. Estrus was defined when a doe accepted mounting by the bucks equipped with canvas aprons. Each doe in estrus got the first transrectal ultrasonography at 24 hours after estrus onset and then every 4 hours until all the preovulatory follicles ovulated. Estrus onset for the TRE and CON groups was 40.3 ± 17.4 (mean ± standard deviation) and 43.3 ± 12.2 hours (P = 0.72), respectively. Estrus duration for the same groups was 28.6 ± 5.4 and 36.7 ± 5.3 hours (P = 0.02), respectively. The mean ovulation time for the TRE and CON groups was 31.4 ± 2.2 and 35.7 ± 3.7 hours (P = 0.04), respectively. The proportion of ovulations that occurred after the end of estrus in the TRE group was higher than in the CON group (86% vs. 33%, respectively; P = 0.05). The number of ovulations for the TRE group was 2.1 ± 0.7; for the CON group, there were 2.2 ± 0.5 ovulations (P = 0.92). It was concluded that copulation by a buck at the beginning of estrus reduced estrus duration and hastened the ovulation time in Boer goats.

A 3-year-old Longhorn heifer was referred to the Veterinary Medical Teaching Hospital of Texas A&M University for inability to get pregnant. Physical examination revealed a small-sized female for age and breed with a normal vulva, vaginal length, and external cervical os. Further assessment by per rectum palpation and trans-rectal ultrasonography revealed a small uterine cervix and cord-like uterine horns with no identifiable ovaries. Additional evaluation including laparoscopy, hormonal evaluation, and genetic analysis allowed ruling out conditions commonly associated with a phenotypic female with infantile or underdeveloped reproductive organs such as freemartin, XY gonadal dysgenesis, testicular feminization, and bilateral ovarian agenesis. Laparoscopy confirmed the presence of a small cervix with small uterine horns and absence of ovaries. Testosterone, progesterone, and 17-β estradiol concentrations were 200.0pg/mL, 1.48ng/mL, and undetectable, respectively. Genetic evaluation determined that the karyotype was 59,X non-mosaic. Evaluation of phenotypically female cattle with infertility and infantile genital organs and absence of ovaries should include cytogenetic analysis to test for possible X monosomy. The 59,X condition should be considered in the differential diagnoses together with freemartin, dysgenesis XY, testicular feminization, and bilateral ovarian agenesis.

The objective was to estimate the effect of palpation per rectum (for early pregnancy diagnosis) on embryo/fetal viability in dairy cattle. A controlled, randomized block-design experiment with two blocks, one by category, and the other by number of embryos, was conducted. Five-hundred-and-twenty pregnant dairy cows and heifers with a viable embryo detected by transrectal ultrasonography (TRUS) between days 29 and 32 after AI were included. The pregnant females were randomly allocated into two nearly equal groups: palpation per rectum (PAL group; n=258) and no palpation per rectum (NPAL group; n=262). The PAL group was submitted to palpation per rectum (PPR) using the fetal membrane slip (FMS) technique once between days 34 and 41 of pregnancy. The fetal membrane slip consisted of compressing the pregnant uterine horn and allowing the chorioallantoic membrane to slip between the fingers. Both groups were submitted to two additional TRUS at days 45 and 60 of pregnancy, to monitor the potential immediate and delayed deleterious effects of PPR on embryo and fetal viability, respectively. A diagnosis of embryo/fetal death was made when there was no embryo/fetal heart beat or the absence of positive signs of pregnancy in an animal previously diagnosed pregnant, or the presence of signs of embryo/fetal degeneration. The overall rate of embryo/fetal death was 14.0% (73/520). Embryonic death (10%; 52/520) was higher than fetal death (4.5%; 21/468; P0.001). Embryo/fetal mortality was higher in cows (16.4%; 59/360) than in heifers (8.8%; 14/160; P0.025) and in cattle with twin (25.5%; 12/47) versus singleton pregnancies (12.9%; 61/473; P0.025), but was not different (P0.05) between PAL (14.7%; 38/258) and NPAL (13.4%; 35/262). In conclusion, PPR between days 34 and 41 of pregnancy using the fetal membrane slip technique did not affect embryo/fetal viability.

The objectives of the present study were to assess the effect of permanent contact of teasers without copulation on the interval from controlled internal drug release (CIDR) removal to estrus onset, estrus duration, ovulation time, number of ovulations, and interval from CIDR removal to ovulation time on estrus-synchronized Boer goats. During the fall season, a controlled randomized design experiment with two groups, control (CON; n = 18) and treatment (TRE; n = 18), was performed. The TRE group was maintained permanently in a pen with an aproned buck immediately after CIDR removal. The CON group was maintained in a different pen without permanent exposure to the male. All females were estrus synchronized with CIDR maintained in the vagina for 7 days and received 50 μg of GnRH im at device insertion and 5 mg of natural prostaglandin F-2α at device removal. Females were considered to be in estrus when they accepted mounting by the aproned bucks. Estrus was detected four times a day after CIDR removal (at 6 AM, 12 noon, 6 PM, and 12 midnight) using bucks with canvas apron as teasers. The ovulation time and number of ovulations were assessed by transrectal ultrasonography starting 24 hours after estrus onset and repeated every 6 hours until complete ovulation was detected. The estrus onset for the CON group was 44.0 ± 8.3 hours and for the TRE group, it was 37.0 ± 7.7 hours (P = 0.01). Estrus duration from the CON group was 43.7 ± 9.2 hours and for the TRE group, it was 38.3 ± 6.6 hours (P = 0.05). The first, last, and mean ovulation times for the CON group were 32.4 ± 5.3, 38.4 ± 3.4, and 35.4 ± 3.9 hours, and for the TRE group, the times were 31.8 ± 2.8, 36.7 ± 3.0, and 35.8 ± 3.6 hours, respectively (P = 0.85, P = 0.23, and P = 0.82, respectively). The number of ovulations for the CON and TRE groups was 2.6 ± 0.7 and 2.6 ± 0.6 ovulations, respectively (P = 0.96). The interval time for CIDR removal to ovulation for the CON group was 79.2 ± 8.2 hours and for the TRE group, the interval time was 73.2 ± 6.2 hours (P = 0.05). It was concluded that the permanent presence of male without copulation with estrus-synchronized does hastened estrus onset, reduced estrus duration, and decreased the interval time from CIDR removal to ovulation without modification of ovulation time and number of ovulations in Boer goats.

Somatic cell nuclear transfer (NT) in cattle is an inefficient process, whereby the production of calves is hindered by low pregnancy rates as well as fetal and placental abnormalities. Interspecies models have been previously used to facilitate the identification of single nucleotide polymorphisms (SNPs) within coding regions of genes to discriminate between parental alleles in the offspring. Here we report the use of a bovine interspecies model (Bos gaurus x Bos taurus) for the assessment and characterization of epigenetic modifications and genomic imprinting in Day 40-old female NT-derived fetuses and placenta. Analysis of NT and control pregnancies indicated disruption of genomic imprinting at the X inactivation-specific transcript (XIST) locus in the chorion, but not the fetus of clones, whereas proper allelic expression of the insulin-like growth factor II (IGF2) and gene trap locus 2 (GTL2) loci was maintained in both the fetus and placenta. Analysis of the XIST differentially methylated region (DMR) in clones indicated normal patterns of methylation; however, bisulfite sequencing of the satellite I repeat element and epidermal cytokeratin promoter indicated hypermethylation in the chorion of clones when compared with controls. No differences were detected in methylation levels in the fetus proper. These results indicate that the nuclear transfer process affects gene expression patterns in the trophectoderm- and inner cell mass-derived tissues to different extents.

The relationship among sperm DNA assays in bulls with different sperm motility and morphology measures has not been reported. The objectives of the present study were to (1) describe Comet assay measures and examine their repeatability (inter- and intra-assay); (2) compare sperm DNA quality assays (i.e., Sperm Chromatin Structure Assay-SCSA; alkaline and neutral Comet assays and Sperm Bos Halomax assay-SBH) in two groups of bulls selected on either greater and lesser sperm motility and morphology (greater compared with lesser); (3) determine the relationship among DNA assays and sperm motility and morphology values. Inter-assay repeatability was greater for the neutral Comet assay as compared to the alkaline Comet assay. Intra-assay repeatability was greater than inter-assay repeatability for both Comet assays. Comet assay dimension measures and percentage tail DNA were the most repeatable for both Comet assays. Among sperm DNA quality assays, only SCSA measures and neutral Comet assay Ghosts (% Ghosts), head diameter and area, and comet area were different between greater and lesser sperm quality groups (P

The use of ultrasonography for pregnancy diagnosis and reproductive tract evaluation in the goat has become more common in the past decade. Pregnancy-specific hormone assays are commercially available for pregnancy determination in goats. Hormonal methods of synchronization of estrus for artificial insemination have been refined, but a number of factors still make pregnancy results variable. Caprine embryo transfer is widely used commercially. More advanced reproductive techniques such as in-vitro production of embryos and cloning have been accomplished in goats; success rates with these techniques will likely rapidly improve.

The objective of this experiment was to compare the effectiveness of two bulls and two different semen treatments on rates of in-vitro fertilization of in-vitro matured bovine oocytes, cleavage and development of embryos to the stage of morula and blastocyst by using a 2 × 2 factorial design. Oocytes (n=946) aspirated from follicles 2-6 mm in diameter were incubated for 20-22 h in TCM-199 supplemented with 5% calf serum, and then inseminated with frozen-thawed semen from two single ejaculates of two Holstein bulls of proven fertility. For the conventional washing method (C), semen was washed twice by centrifugation at 500 g for 5 min with modified BO medium (BO: without glucose or bovine serum albumin (BSA)) supplemented with 10 mM caffeine and 2.5 IU/ml heparin (BO-wash), and then diluted with the same amount of BO containing 20 mg/ml BSA (BO-dilute). For the Percoll washing method (P) semen was layered on a 45 to 90% percoll gradient in a 15 ml centrifugation tube. After 30 min of centrifugation at 700 g, the sperm pellet was recovered from the bottom of the tube, and then resuspended in BO-wash and double diluted with BO-dilute. With this sperm suspension insemination drops (100 μl) were prepared and the sperm was cultured with oocytes for 5 h. Then the oocytes were transferred to CR1aa supplemented with 5% calf serum for 7 days. The rates of cleavage and morula or blastocyst development of oocytes fertilized in vitro were similar in P (14.2% and 7.9%, respectively) and C (17.7% and 7.7%, respectively) methods for bull A, but these rates were significantly higher (P

Objective—To determine effects of per rectal amniotic sac palpation (ASP) for pregnancy diagnosis during early gestation on pregnancy loss in lactating cows. Design—Controlled, randomized block design. Animals—368 pregnant dairy cows. Procedures—Pregnancy was detected via transrectal ultrasonography (TRUS) at day 29 (day of estrus = day 0), and cows were allocated into a control group (n = 167 cows) and ASP group (180). Control cows were not subjected to pregnancy diagnosis via palpation per rectum. Per rectal ASP was performed between days 34 and 43 by only 1 experienced veterinarian. All cows were reevaluated with TRUS on days 45, 60, and 90. Results—21 cows were removed because of illness. Pregnancy loss between days 29 and 90 occurred in 44 of 347 (12.7%) cows. Pregnancy loss for the control and ASP groups from days 29 to 90 occurred in 22 of 167 (13.2%) and 22 of 180 (12.2%) cows, respectively. Late embryonic pregnancy loss (days 29 to 45) for the control and ASP groups occurred in 18 (10.8%) and 15 (8.3%) cows, respectively. Early fetal pregnancy loss (days 46 to 60) for the control and ASP groups occurred in 2 of 149 (1.3%) and 6 of 165 (3.6%) cows, respectively, and late fetal pregnancy loss (days 61 to 90) for the same groups occurred in 2 of 147 (1.4%) and 1 of 159 (0.6%) cows, respectively. Conclusions and Clinical Relevance—Pregnancy diagnosis via per rectal ASP during early gestation did not increase pregnancy loss in dairy cattle.

Objective—To determine the effect of palpation per rectum (PPR) by use of 1 or 2 fetal membrane slips (FMSs) for pregnancy diagnosis during early gestation on pregnancy loss in dairy cattle. Design—Controlled, randomized block design. Animals—928 healthy pregnant cattle. Procedures—All cattle were determined to be pregnant by use of transrectal ultrasonography at approximately day 31 after estrus and randomly allocated into 2 groups (control group [n = 476 cows] and palpation group [452]). The control group was not subjected to pregnancy diagnosis via PPR. The palpation group was subdivided into 2 groups (PPR FMS 1 [n = 230 cows] and PPR FMS 2 [222]), which involved PPR and pregnancy diagnosis via 1 or 2 FMSs, respectively, during the same examination, which was performed by 1 veterinarian between days 34 and 43 after estrus. All cattle were reevaluated by use of transrectal ultrasonography on days 45 and 60 to determine viability of the embryo and fetus, respectively. Results—Overall pregnancy loss between days 31 and 60 was 14.1%. Pregnancy loss for the control, PPR FMS 1, and PPR FMS 2 groups from days 31 to 60 was 14.5%, 12.6%, and 14.9%, respectively. Embryonic pregnancy loss for the control, PPR FMS 1, and PPR FMS 2 groups was 12.4%, 9.1%, and 9.5%, respectively. Fetal pregnancy loss for the same groups was 2.4%, 3.8%, and 5.9%, respectively. Conclusions and Clinical Relevance—Pregnancy diagnosis via 1 or 2 FMSs performed during PPR in early gestation did not increase pregnancy loss in dairy cattle.

The objective was to evaluate the accuracy of an ELISA for pregnancy specific protein B (PSP-B) for early pregnancy diagnosis in dairy cattle. Blood from lactating (>100 d postpartum) dairy cows (n = 738), was collected on Days 28, 30, and 35 (Day 0 = estrus), analyzed with an ELISA for PSP-B, and the cows designated as pregnant, probable, unlikely, or non-pregnant. Immediately after blood collection, transrectal ultrasonography (TRUS) was done for pregnancy diagnosis, and the results used as a criterion standard test for comparison with PSP-B. At Day 28, 46.3% were diagnosed by TRUS as pregnant. The PSP-B sensitivity was 93.9% on Day 28 and similar on Days 30 and 35. The PSP-B specificity, positive predictive value, negative predictive value, and accuracy were all >94% on Day 28 and similar on Days 30 and 35. However, the accuracy was significantly less compared to TRUS (P < 0.01). The percentage of all samples from cows that were probably pregnant or unlikely to be pregnant was 5.6%. At Days 28, 30, and 35, percentages of uncertain samples were 8.5, 4.8, and 3.3%, respectively (P < 0.01), and Kappa values were 0.92, 0.92, and 0.95. False negative and false positive results were attributed to low concentrations of PSP-B in pregnant animals and to persistence of pregnant concentrations of PSP-B in females with pregnancy loss, respectively. In conclusion, PSP-B ELISA was a sensitive, specific, and accurate test for pregnancy diagnosis (relative to TRUS) at Days 28, 30, and 35 after breeding.

Cloning using somatic cell nuclear transfer (SCNT) may be a useful tool for conserving genetic diversity and for propagating exotic and/or endangered animal species. Somatic cells can be obtained easily, expanded in culture, cryopreserved, and thawed at a later date for use in NT. Significant challenges relevant to using SCNT for cloning wild and endangered animal species include the need for using interspecies NT and interspecies embryo transfer. Animal care and welfare issues raised that are unique to exotic and endangered species also are raised. In this chapter, the methods used in attempts to clone the wild animal species of Desert Bighorn Sheep are described.

The objective of the present study was to determine differences in time of detection of pregnancy between heifers and cows and the interval after insemination at which the maximum sensitivity and negative predictive value of transrectal ultrasonography were obtained. One-thousand-four-hundred transrectal ultrasonographies (TRUS-1; 1,079 in cows and 321 in heifers) were performed using a 5-MHz linear-array transducer. The cattle were randomly assigned to have TRUS performed once between days 24 and 30 (estrus=day 0) in cows or between days 21 and 27 in heifers. Every TRUS diagnosis was subsequently compared with a second TRUS diagnosis (TRUS-2), performed 3-8 days later, after day 30 (range 31-38) for cows and after day 27 (range 28-35) for heifers. The sensitivity and specificity between cows and heifers for the common days of TRUS (from 24 to 27) were compared. In cows, sensitivity increased gradually from 74.5% at day 24 to 100% at day 29 (P0.01). Specificity increased from days 24-25 and reached a plateau of 96.6% on day 26 (P0.01). In heifers, sensitivity increased from 50% at day 21 to 100% at day 26 (P0.01). Specificity increased from 87.5% at day 21 and remained steady at 94% starting on day 23 (P0.05). The sensitivity for cows and heifers was 89.2 and 96.8%, respectively (P0.05) and the specificity was 93.0 and 93.4% (P0.05). In this study, heifers were diagnosed pregnant earlier than cows, and the maximum sensitivity and negative predictive value were obtained 3 days earlier in heifers than cows (days 26 and 29, respectively).

Immunomagnetic bead separation coupled with bead beating and real-time PCR was found to be a very effective procedure for the isolation, separation, and detection of Mycobacterium avium subsp. paratuberculosis from milk and/or fecal samples from cattle and American bison. Samples were spiked with M. avium subsp. paratuberculosis organisms, which bound to immunomagnetic beads and were subsequently lysed by bead beating; then protein and cellular contaminants were removed by phenol-chloroform-isopropanol extraction prior to DNA precipitation. DNA purified by this sequence of procedures was then analyzed by conventional and real-time IS 900 -based PCR in order to detect M. avium subsp. paratuberculosis in feces and milk. By use of this simple and rapid technique, 10 or fewer M. avium subsp. paratuberculosis organisms were consistently detected in milk (2-ml) and fecal (200-mg) samples, making this sensitive procedure very useful and cost-effective for the diagnosis of clinical and subclinical Johne's disease (paratuberculosis) compared to bacteriological culture, which is constrained by time, labor, and expense under diagnostic laboratory conditions.

To date, the efficiency of pig cloning by nuclear transfer of somatic cell nuclei has been extremely low, with less than 1% of transferred embryos surviving to term. Even the utilization of complex procedures such as two rounds of nuclear transfer has not resulted in greater overall efficiencies. As a result, the applicability of the technology for the generation of transgenic and cloned animals has not moved forward rapidly. We report here a simple nuclear transfer protocol, utilizing commercially available in vitro-matured oocytes, that results in greater than 5% overall cloning efficiency. Of five recipients receiving nuclear transfer embryos produced with a fetal fibroblast cell line as nuclear donor, all five established pregnancies by day 28 (100%), and 4/5 (80%) went to term. Efficiencies for each transfer were 7% (9 piglets/128 doublets transferred), 5% (5/100), 12% (7/59), and 6.6% (7/106). The overall efficiency in all recipients was 5.5% and in pregnant recipients 7.7%, with a total of 28 cloned piglets produced. With the average fusion rate being 58%, the percentage of fused doublets producing a live piglet approached 12%. The method described here can be undertaken by a single micromanipulator at a reasonable cost, and should facilitate the broad utilization of porcine cloning technology in transgenic and nontransgenic applications.

Juan E. Romano *, James A. Thompson , David W. Forrest , Mark E. Westhusin , Michael A. Tomaszweski , Duane C. Kraemer b,c a Large Animal Clinical Sciences. College of Veterinary Medicine, Texas A&M University, College Station, TX 77845, USA b Veterinary Physiology and Pharmacology, College of Veterinary Medicine, Texas A&M University, College Station, TX 77845, USA c Department of Animal Science, College of Agriculture and Life Sciences, Texas A&M University, College Station, TX 77845, USA

The methods for collecting and freezing deer semen have been, for the most part, limited to two species; red deer ( Cervus elaphus) and fallow deer ( Dama dama) (Asher et al., 2000 Anim. Reprod. Sci. 62, 195–211). The object of this study was to evaluate the progressive motility and effects of a thermal stress test on white-tailed deer (WTD) semen frozen in Biladyl extender (Mini Tube, Verona, WI, USA). Semen was collected by electroejaculation from WTD bucks (n=7, ages 1.5–2.5 years) during the breeding season. This trial was the second collection for one buck (#0025) and the third collection for the other 6 bucks. The bucks were immobilized with a xylazine/ketamine mixture i.m. (2mgkg−1 Vedco, Inc., St. Joseph, MO, 2.2mgkg−1 ketamine HCl, Fort Dodge Animal Health, Fort Dodge, IA, USA) and electroejaculated with a Pulsator IV unit (Lane Manufacturing, Denver, Co). Semen was extended 1:1 with Biladyl A, and then slowly cooled to 4°C. Once cooled, semen was extended with equal amounts of Biladyl part A, then part B, to a final concentration of 160×106cells/mL. The extended semen was then loaded into 0.25-cc straws, placed over liquid nitrogen (LN2) in vapors (−80°C) for 10min, and then plunged into LN2. Straws were stored in a LN2 tank for 3 months. Semen was thawed in a 38.5°C water bath for 45s, then placed in a warm test tube and incubated at 38.5°C for 5min before progressive motility was evaluated using a computer program (Sperm Vision, Mini Tube). A thermal stress test was performed by incubating thawed samples at 38.5°C for 1h. Results of the stress test were graded as either passed (progressive motility ≥50%) or failed (progressive motility