Pre- and Post-partum Concentrations of Interleukin 1α, Interleukin 8, and α1-Acid Glycoprotein in Vaginal Fornix and Endometrium of Dairy Cows With Clinical Cervicitis.

Innate immunity is the principal sensor responsible of the local immune response to control mucosal bacterial contamination of the reproductive tract after parturition, triggering a pro-inflammatory process in the mucosa of the uterus, the vaginal and the cervix. However, knowledge about the inflammation process and outcome of the cervix in dairy cows is scarce even though it plays an important anatomic and functional role between the vagina and the uterus. The objective of the present study was to describe the cellular and humoral local innate immune response during clinical cervicitis (CC) in the uterus and vaginal fornix in pre- and post-partum periods of dairy cows. A retrospective descriptive study was performed involving 26 animals, characterized as clinical cervicitis cows (n = 19) and healthy cows (n = 7). Blood and mucus of the different compartments of the genital tract were sampled and records of the cows' genital exam were performed four times: -1 w (day -7 ± 2, prepartum), +1 w (day +7 ± 4), +3 w (day +21 ± 4) and +5 w (day +35 ± 4) postpartum. Clinical cervicitis was defined as cows exhibiting a cervix grade-2 and healthy cows were defined as a cow clinically normal with a grade-0 cervix at time +5 w. Blood white cell count, vaginal fornix and endometrial neutrophils percentage, and the concentrations of interleukin 1α (IL1), interleukin 8 (IL8), and α1-acid glycoprotein (AGP) in mucus were determined. The results showed that 23% of the cows were categorized as CC at time +5 w. Cases of CC with purulent vaginal discharge or subclinical endometritis shown the highest cytokine production. At +3 w, IL1, IL8, and AGP concentrations in the uterus and the fornix were significantly higher in CC than healthy cows (CH). In conclusion, the 3-week postpartum is a critical point to evaluate cytokines and acute phase proteins; where IL1 and IL8 variation kept a direct relation with neutrophils numbers and function. The presence of AGP in the endometrium infer a homeostatic proinflammatory protective balance effect, modulating the local uterine innate immune response during peripartum.

Outcome Following Partial Unilateral Hysterectomy by Standing Flank Approach in Dairy Cattle.

OBJECTIVE: To describe the clinical findings, surgical treatment, and long-term outcome of dairy cattle undergoing partial hysterectomy with or without unilateral ovariectomy. STUDY DESIGN: Retrospective case series. ANIMALS: Dairy cattle (n = 7). METHODS: Medical records (June 2007-June 2014) of dairy cattle that had partial hysterectomy with or without ipsilateral ovariectomy were reviewed. Follow-up data were obtained by telephone interviews with owners and referring veterinarians between 6 months and 7 years following discharge. RESULTS: Seven dairy cattle met the inclusion criteria. Diagnoses made intra- or postoperatively were ovarian abscess with multiorgan adhesions (4 cows), segmental aplasia of the uterus (2 cows), and uterine lymphosarcoma (1 cow). Partial hysterectomy with or without unilateral ovariectomy was performed without surgical complications by standing flank approach. All cows were successfully discharged from the hospital (short-term prognosis). Five cows became pregnant, and 4 delivered live calves and had a satisfactory productive life without long-term complications. CONCLUSION: Partial hysterectomy by standing flank approach should be considered as a viable treatment option for unilateral ovarian or uterine problems in dairy cattle. Cattle undergoing partial hysterectomy with or without ipsilateral ovariectomy are capable of satisfactory reproduction and milk production after surgery with no short- or long-term complications.

Fetal mummification in the major domestic species: current perspectives on causes and management.

Fetal mummification is an uncommon condition in most domestic species. While most often seen in multiparous and polytocous species like swine, it is also observed in monotocous species when the fetus is retained for a long time. The low prevalence of the condition may help explain the scarcity of information in the literature. To further complicate the study of this phenomenon, the physiological mechanisms that maintain pregnancy vary between species, implying different pathways for the condition. The exact outcome of early fetal mortality is unpredictable, and is influenced by several factors, including the cause of fetal mortality, differences in pregnancy between species, stage of gestation at fetal death, and number of fetuses. Based on our current knowledge of natural fetal mummification events, there are a number of prerequisites for the process of fetal mummification to occur. Examining the circumstances associated with fetal mummification can help scientists better understand the etiology and clinical situation in different species. The objective of this article is to review fetal mummification in the major domestic species: cattle, goats, sheep, horses, swine, dogs, and cats. This paper discusses the clinical situation, the most common and important etiologies, and the treatment approaches for restoring future pregnancy in the female, and where applicable, herd fertility.

Effect of superovulation on uterine and serum biochemical parameters and its potential association with transferable embryos in Holstein dairy cows.

The objective of this study was to determine the effects of superovulation (SOV) on serum and uterine biochemical parameters, uterine bacteriology and cytology and number of transferable embryos (TE). Dairy cows were placed on a Presynch/CIDR Synch protocol. The SOV group was superovulated, induced in estrus, and inseminated, whereas the control group was induced in estrus and inseminated without SOV. Uterine bacteriology and cytology and uterine and serum biochemical parameters were measured at day 7 of the estrous cycle to start the SOV protocol, as well as on the day of embryo recovery (DER). The SOV group produced 7.5 ± 6.7 oocytes/embryos, of which 3.4 ± 4.7 were TE. Serum urea and E2 and uterine Glu, CK, LDH, TP, P4 and PGFM in the control group and serum P4 and PGFM and uterine LDH and PGFM in the SOV group were significantly higher (p < 0.01) at DER than day 7. At DER, uterine urea, LDH, PGFM and TP and serum urea, LDH, PGFM, and P4 concentrations were higher (p < 0.01) in the SOV group than the control. There was no significant variation in uterine bacteriology or cytology. Overall, these results infer that SOV affects both serum profile and uterine secretions, and that these changes may influence the number of TE.

Identification of single nucleotide polymorphisms in the bovine follicle-stimulating hormone receptor and effects of genotypes on superovulatory response traits.

In dairy cows, there is evidence that failure to respond to superovulation protocols is a heritable trait. In women, genotyping for the p.N680S single nucleotide polymorphism (SNP) in the follicle-stimulating hormone receptor (FSHR) gene may help identify poor responders before ovarian stimulation is initiated. Our objectives were to identify SNPs in the coding region of the bovine FSHR gene and to investigate the effect of FSHR genotypes on superovulatory response in Holstein cattle. Sequencing of FSHR exons 1-10 revealed seven SNPs. Three were non-synonymous mutations (c.337C>G, c.871A>G and c.1973C>G). SNP c.337C>G encodes for a proline-to-alanine (p.Pro113Ala) amino acid replacement in the extracellular ligand-binding domain of the receptor. PCR-RFLP analyses showed that homozygous GG Holstein cows present a higher percentage of viable embryos, whereas GG and CG animals have less unfertilised oocytes. SNP c.871A>G results in an isoleucine-to-valine (p.Ile291Val) modification, and homozygous AA animals present lower embryo yield after superovulatory treatments. SNP c.1973C>G corresponds to a threonine-to-serine (p.The658Ser) modification in the intracellular carboxyl-terminal domain of the FSHR protein, and homozygous GG Holstein cows were associated with a lower embryo yield and a higher percentage of unfertilised oocytes. Our results suggest that specific alleles of the bovine FSHR gene are associated with variations in embryo yield and in the number of unfertilised oocytes.

Therapeutic efficiency of antibiotics and prostaglandin F2α in postpartum dairy cows with clinical endometritis: an evidence-based evaluation.

The occurrence of vaginal discharge in postpartum dairy cows is generally diagnosed as clinical endometritis. This uterine condition is associated with reduced fertility and economic loss for the dairy industry. Therapeutic approaches include the systemic or intrauterine application of antibiotics or the injection of prostaglandin F2α and analogues to cause luteolysis and uterine contractions to evacuate the infected content. The treatment of clinical endometritis remains a subject of considerable controversy in the literature. Better understanding of the reproductive biology of normal versus abnormal uterine involution and immune mechanisms will allow more efficient diagnostic methods and a more efficient therapeutic approach.

Expression of steroidogenic proteins in bovine placenta during the first half of gestation.

The aim of the present study was to determine the occurrence and localisation of the principal steroidogenic proteins in bovine placenta from Day 50 to Day 120 of pregnancy. Immunohistochemistry revealed that, at all stages investigated, bovine steroidogenic acute regulatory protein (StAR), cytochrome P45011A1 and hydroxy-δ-5-steroid dehydrogenase, 3ß- and steroid δ-isomerase 1 proteins were found principally at the fetomaternal interdigitations: the chorionic villus and maternal septum. Moreover, caruncular epithelial cells and uninucleate trophoblast cells were the principal cells detected that were positive for the three markers. Western blot analysis showed that only caruncular tissue expressed all three steroidogenic markers; in contrast, cotyledons only expressed StAR and cytochrome P45011A1. Immunoblot results showed a complementary pattern of StAR and cytochrome P45011A1 expression between caruncles and cotyledons at different stages. These observations suggest that, in early pregnancy, the maternal compartment contributes significantly to bovine placental steroidogenesis, particularly for the synthesis of progesterone. Furthermore, the variation in StAR and cytochrome P45011A1 expression between caruncular and cotyledonary tissues across gestation suggests that placental steroidogenesis requires cell-to-cell communication between maternal and fetal cells.

Effect of reproductive disorders and parity on repeat breeder status and culling of dairy cows in Quebec.

This study quantified the effect of peripartum reproductive disorders and parity on repeat breeder status and involuntary culling of dairy cows. Reproductive data of 418 383 lactations were taken from a computerized databank of health records for dairy cows. A logistic regression model was used with dystocia, retained placenta (RP), metritis complex, and parity as fixed effect risk factors and herd entered as the random effect. Of the peripartum problems studied, dystocia had the greatest effect on future fertility. Dystocia increased the odds of a cow being a repeat breeder by 44% [odds ratio (OR): 1.44; confidence interval (CI): 1.37 to 1.51]. Compared to first-parity cows, cows in second, third, and fourth parities had significantly higher odds of being a repeat breeder: 18% (OR: 1.18; CI: 1.16 to 1.20); 24% (OR: 1.24; CI: 1.21 to 1.26); and 42% (OR: 1.42; CI: 1.39 to 1.45), respectively. The odds for second-, third-, or fourth-parity repeat breeders being culled were 24% (OR: 1.24; CI: 1.20 to 1.28); 39% (OR: 1.39; CI: 1.35 to 1.43); and 67% (OR: 1.67; CI: 1.62 to 1.71) respectively, while peripartum reproductive problems had less of an effect.

Assessment of fetal well-being in cattle by ultrasonography in normal, high-risk, and cloned pregnancies.

This study determined ultrasonographic parameters of fetuses and uterine adnexa in late pregnancy in normal, cloned, and high-risk pregnancies in relation to perinatal and neonatal outcome. Ten cows with normal pregnancies (CONTROL, mean pregnancy length 273 d), 10 sick cows with potentially compromised pregnancies (HIGH-RISK, mean pregnancy length 267 d), and 10 heifers with cloned pregnancies (CLONED, mean pregnancy length 274 d) were examined at more than 260 d of gestation. There was no difference in mean fetal heart rates among the groups. The cloned calves were heavier (57 ± 8 kg) than calves from CONTROL group (36 ± 7 kg), and calves from HIGH-RISK group (37 ± 13 kg) (P = 0.003). The diameter of the thoracic aorta was positively correlated (R = 0.62) with fetal birth weight in the CONTROL group (P = 0.01). Fetal activity was not associated with survival. The results suggest that transabdominal ultrasonographic assessment of the fetal well-being may serve as a potential tool for evaluation of the fetoplacental unit.

Ultrasound imaging of the bull reproductive tract: an important field of expertise for veterinarians.

Diagnosing male reproductive system pathologies can often be frustrating because of the challenge involved in precisely determining their site, severity, and prognosis. The introduction of complementary ultrasonographic examination enables clinicians to address these important questions. This procedure should be performed not only on bulls destined to artificial insemination, but on all farm bulls. The examination is easy to perform with a versatile ultrasonographic unit designed for bovine theriogenology. To recognize abnormal tissues, however, the operator must have an excellent knowledge of the ultrasonographic anatomy of the reproductive system. This article discusses the basis of ultrasound technique for male reproductive tract examination. Ultrasound evaluation of physiologic and pathologic conditions of external and internal reproduction organs is proposed.

Ultrasonographic fetal well-being assessment, neonatal and postpartum findings of cloned pregnancies in cattle: a preliminary study on 10 fetuses and calves.

Cloned pregnancies in cattle are considered to be at risk due to a variety of fetal or adnexal abnormalities. Data is lacking concerning the possibility of transabdominal ultrasonography in the assessment of these high risk pregnancies. Transabdominal ultrasonography has rarely been reported in the assessment of bovine cloned pregnancies. Ten Holstein heifers carrying 8-month-old cloned fetuses were assessed by transabdominal ultrasonographic examination during the 3rd trimester of pregnancy. Fetal heart rates (FHR), movements, adnexal appearance, and placentome size were recorded. The outcome of the pregnancies was also noted and potential indicators of fetal demise recorded. Survival rate 1 week after birth was 30%. Mean FHR was 113 beats per minute (range: 92 to 128 bpm) during the fetal ultrasonography. No correlation between FHR and fetal activity was found. Fetal hyperactivity and imaging of hyperechoic particles in both allantoic and amniotic fluids were possible signs of fetal distress. Despite the size of the fetus and the deep bovine abdomen, fetal transabdominal ultrasonography can be performed in cattle. This preliminary study points to the necessity of further larger studies for defining normal and abnormal findings in bovine late pregnancy.

Retrospective case study of fetal mummification in cows that did not respond to prostaglandin F2alpha treatment.

Mummification of bovine fetuses is an uncommon condition, and cows do not always respond to treatment with prostaglandin F2alpha. The objective of the present retrospective and descriptive case study was to determine the conception rate and survival time of nonresponsive, prostaglandin F2alpha (PGF2alpha)-treated cows (n = 14), following hysterotomy or medical treatment and manual removal. Animal records from 1990 to 2005 from the Centre Hospitalier Universitaire Vétérinaire (CHUV) of the Université de Montréal were studied. Inclusion criteria were the nonexpulsion of the mummified fetus following PF2alpha treatment and absence of concomitant conditions upon physical examination. Of the animals included in the study, 36% (n = 5) became pregnant after extraction of the mummified fetus by hysterotomy and 0% conceived after medical treatment and manual extraction. In this study, hysterotomy represented an effective approach for extracting mummified fetuses from cows that did not respond to PF2alpha treatment.

In vitro culture and somatic cell nuclear transfer affect imprinting of SNRPN gene in pre- and post-implantation stages of development in cattle.

BACKGROUND: Embryo in vitro manipulations during early development are thought to increase mortality by altering the epigenetic regulation of some imprinted genes. Using a bovine interspecies model with a single nucleotide polymorphism, we assessed the imprinting status of the small nuclear ribonucleoprotein polypeptide N (SNRPN) gene in bovine embryos produced by artificial insemination (AI), in vitro culture (IVF) and somatic cell nuclear transfer (SCNT) and correlated allelic expression with the DNA methylation patterns of a differentially methylated region (DMR) located on the SNRPN promoter. RESULTS: In the AI group, SNRPN maternal expression is silenced at day 17 and 40 of development and a third of the alleles analyzed are methylated in the DMR. In the IVF group, maternal transcripts were identified at day 17 but methylation levels were similar to the AI group. However, day-40 fetuses in the IVF group showed significantly less methylation when compared to the AI group and SNRPN expression was mostly paternal in all fetal tissues studied, except in placenta. Finally, the SCNT group presented severe loss of DMR methylation in both day-17 embryos and 40 fetuses and biallelic expression was observed in all stages and tissues analyzed. CONCLUSION: Together these results suggest that artificial reproductive techniques, such as prolonged in vitro culture and SCNT, lead to abnormal reprogramming of imprinting of SNRPN gene by altering methylation levels at this locus.

Twin pregnancy experimental model for transvaginal ultrasound-guided twin reduction in mares.

Multiple pregnancies are still an important cause of noninfectious abortion, stillbirth, neonatal mortality, and significant delays in reproductive performance in mares. Despite new management techniques, reduction in multiple pregnancies is an ongoing preoccupation and challenge for the equine veterinarian. The aim of the present study was to establish a twin pregnancy experimental model in the mare to study the effectiveness of a transvaginal ultrasound-guided embryonic vesicle injection. Mares in heat were inseminated and then received an embryo at day 7 of the estrous cycle. At days 14 and 30, 53.5% (n = 23) and 23% (n = 10) of the mares, respectively, were carrying twins. Twin pregnancies were reduced at day 30 by transvaginal ultrasound-guided puncture of the embryonic vesicle (control, n = 5) or by transvaginal ultrasound-guided injection (TVUEVI) of 25 mg of amikacin into the embryonic vesicle (n = 5). The TVUEVI treatment had a 40% success rate and no significant variations in progesterone and prostaglandin metabolite were observed. Even though the technique does not seem very effective, the experimental model could be useful for clinical research in embryo reduction and early embryonic loss.

Accuracy of a rapid enzyme-linked immunosorbent assay to measure progesterone in mares.

The aim of this study was to validate an enzyme-linked immunosorbent assay (ELISA) for the measurement of progesterone (P4) in mares. Specifically, the objectives were as follows: (1) to determine the specificity and sensitivity of the ELISA test for determination of P4, (2) to measure the potential agreement between the 2 people performing the test, and 3) to evaluate the effect of time on the outcome. Ten mares were sampled on the day before ovulation (D-1), and on days 1 (D1), 3 (D3), and 5 (D5) following ovulation, during the reproductive season. While mares were cycling regularly, estrus was induced by the injection of 5 mg of prostaglandin (PGF2) and monitored starting on the 4th day by daily transrectal palpation and ultrasonography to determine the time of ovulation. Blood was collected and all samples (n=96) were assayed for P4 by a semiquantitative ELISA, by chemiluminescent immunoassay, and by radioimmunoassay (RIA). Based on the RIA, values of P4 on D-1, D1, D3, and D5 were significantly different (P < 0.0001) with mean and standard deviation(s) of 0.004, s = 0.52; 2.05, s = 2.58; 8.37, s = 4.17; and 12.76, s = 4.00 ng/mL respectively. The sensitivity and specificity of the semiquantitative assay were 94% and 95%, respectively for the lowest values of P4 (< 1.0 ng/mL). The value of kappa was 0.90 between 2 individuals performing the test. In conclusion, these results suggest that the semiquantitative test may be used reliably and economically to evaluate P4 levels in equine plasma in the clinical

Fetal well-being assessment in bovine near-term gestations: current knowledge and future perspectives arising from comparative medicine.

Cloning technology is associated with multiple losses throughout pregnancy and in the neonatal period. Any maternal or fetal disease can compromise pregnancy. A paucity of data are available on bovine fetal well-being in late pregnancy; development of well-being assessment methods might augment early diagnosis of abnormal pregnancy or fetal distress, allowing early intervention. This review presents the current knowledge on fetal well-being based on bovine, ovine, equine, and human studies, as well as interesting research parameters that have been studied in other species and not yet investigated in cattle. Transabdominal ultrasonography allows for diagnosis of large placentomes and hydrallantois that frequently accompany clone pregnancies. Fetal inactivity or large hyperechoic particles imaged within the fetal annexes are associated with fetal distress or death, and should be reassessed to confirm compromised pregnancy. Measurements of different fetal parameters (thoracic aorta, metacarpal or metatarsal thickness) could be reliable tools for early detection of the large offspring syndrome commonly found in cloned calves.

Low-density lipoprotein receptor-related protein 8 (LRP8) is upregulated in granulosa cells of bovine dominant follicle: molecular characterization and spatio-temporal expression studies.

The low-density lipoprotein (LDL) receptor-related protein 8 (LRP8) is a member of the LDL receptor family that participates in endocytosis and signal transduction. We cloned the full-length bovine LRP8 cDNA in granulosa cells (GC) of the dominant follicle (DF) as well as several LRP8 mRNA splicing variants, including a variant that contains a proline-rich cytoplasmic insert (A759-K817) that is involved in intracellular signaling. Expression of the A759-K817 variant was analyzed in the GC of follicles at different developmental stages: the small follicle (SF; 2-4 mm), the DF at Day 5 (D5) of the estrus cycle, ovulatory follicles (OF) 24 h after hCG injection, and corpora lutea (CL) at D5. RT-PCR analysis showed that expression was predominant in the GC of DF compared to other follicles and CL (P<0.0001), whereas the expression of other related receptors, such as LDLR and VLDLR, did not show differences. Temporal analyses of follicular walls from the OF following hCG treatment revealed a decrease in LRP8 mRNA expression starting 12 h post-hCG treatment (P<0.0001). LRP8 protein was exclusively localized to the GC, with higher levels in the DF than in the SF (P<0.05). RELN mRNA, which encodes an LRP8 ligand, was highly expressed in the theca of the DF as compared to the OF (P<0.004), whereas MAPK8IP1 mRNA, which encodes an LRP8 intracellular interacting partner, is expressed in the GC of the DF. These results demonstrate the differential expression patterns of LRP8, RELN, and MAPK8IP1 mRNAs during final follicular growth and ovulation, and suggest that a RELN/LRP8/MAPK8IP1 paracrine interaction regulates follicular growth.

Clinical diagnosis of uterine didelphia in Ayrshire heifer.

A 24-month-old Ayrshire heifer was referred because of infertility. A clinical diagnosis of uterus didelphys was established after a complete genital examination before the animal was culled from the herd.

Induction of alpha-caveolin-1 (alphaCAV1) expression in bovine granulosa cells in response to an ovulatory dose of human chorionic gonadotropin.

Caveolins are implicated in endocytosis, cholesterol trafficking and signal transduction. A cDNA fragment corresponding to caveolin-1 (CAV1) was identified in a mRNA profiling expression study in bovine granulosa cells (GC) following human chorionic gonadotropin (hCG)-induced ovulation. Thus, we have characterized CAV1 cDNA and studied its spatio-temporal expression pattern in bovine ovarian follicles. The full-length bovine alphaCAV1 cDNA was cloned and encodes a putative 22 kDa protein. Expression of alphaCAV1 was studied in bovine GC obtained from follicles at different developmental stages: small follicles (SF: 2-4 mm), dominant follicles (DF), ovulatory follicles (OF: 24 hr post-hCG), and corpus luteum (CL). Semiquantitative RT-PCR analysis showed a 6.5-fold increase in alphaCAV1 mRNA in GC of OF versus DF (P < 0.0001), whereas CAV2 mRNA was increased by only twofold (P < 0.0007). Temporal expression of alphaCAV1 mRNA from OF recovered at 0, 6, 12, 18, and 24 hr after hCG injection showed an 8.5-fold increase of alphaCAV1 mRNA after 24 hr compared to 0 hr (P < 0.0018) whereas no significant variation was detected for CAV2. Immunoblot demonstrated an initial increase in alphaCAV1 protein level 12 hr post-hCG, reaching a maximum at 24 hr. Immunohistochemical localization of CAV1 was observed in GC of OF isolated 18 and 24 hr after hCG injection, whereas no signal was detected in GC of DF and SF. The induction of alphaCAV1 in GC of OF suggests that alphaCAV1 likely contributes to control the increase in membrane signaling that occurs at the time of ovulation and luteinization.

Somatic cell nuclear transfer alters peri-implantation trophoblast differentiation in bovine embryos.

Abnormal placental development limits success in ruminant pregnancies derived from somatic cell nuclear transfer (SCNT), due to reduction in placentome number and consequently, maternal/fetal exchange. In the primary stages of an epithelial-chorial association, the maternal/fetal interface is characterized by progressive endometrial invasion by specialized trophoblast binucleate/giant cells (TGC). We hypothesized that dysfunctional placentation in SCNT pregnancies results from aberration in expression of genes known to be necessary for trophoblast proliferation (Mash2), differentiation (Hand1), and function (IFN-tau and PAG-9). We, therefore, compared the expression of these factors in trophoblast from bovine embryos derived from artificial insemination (AI), in vitro fertilization (IVF), and SCNT prior to (day 17) and following (day 40 of gestation) implantation, as well as TGC densities and function. In preimplantation embryos, Mash2 mRNA was more abundant in SCNT embryos compared to AI, while Hand1 was highest in AI and IVF relative to SCNT embryos. IFN-tau mRNA abundance did not differ among groups. PAG-9 mRNA was undetectable in SCNT embryos, present in IVF embryos and highest in AI embryos. In postimplantation pregnancies, SCNT fetal cotyledons displayed higher Mash2 and Hand1 than AI and IVF tissues. Allelic expression of Mash2 was not different among the groups, which suggests that elevated mRNA expression was not due to altered imprinting status of Mash2. The day 40 SCNT cotyledons had the fewest number of TGC compared to IVF and AI controls. Thus, expression of genes critical to normal placental development is altered in SCNT bovine embryos, and this is expected to cause abnormal trophoblast differentiation and contribute to pregnancy loss.