Analysis of the uterine flush fluid proteome of healthy mares and mares with endometritis or fibrotic endometrial degeneration.

The objective of this study was to evaluate the differences in the uterine flush fluid proteome between healthy mares and mares with endometritis or fibrotic endometrial degeneration (FED). Uterine flush fluid samples were collected from healthy mares (n=8; oestrus n=5 and dioestrus n=3) and mares with endometritis (n=23; oestrus n=14 and dioestrus n=9) or FED (n=7; oestrus n=6 and dioestrus n=1). Proteomic analysis was performed using label-free liquid chromatography-tandem mass spectrometry. Of 216 proteins identified during oestrus, 127 were common to all three groups, one protein was exclusively detected in healthy mares, 47 proteins were exclusively detected in mares with endometritis and four proteins were exclusively detected in mares with FED. Of 188 proteins identified during dioestrus, 113 proteins were common between healthy mares and mares with endometritis, eight proteins were exclusively detected in healthy mares and 67 proteins were exclusively detected in mares with endometritis. Quantitative analysis revealed a subset of proteins differing in abundance between the three groups during oestrus and between healthy mares and mares with endometritis during dioestrus. These results provide a springboard for evaluation of specific proteins as biomarkers of uterine health and disease and for investigation of their roles in the establishment and maintenance of pregnancy.

Qualitative analysis and functional classification of the uterine proteome of mares in oestrus and dioestrus.

Quantitative analysis of the uterine flush fluid proteome of mares in oestrus and dioestrus has been previously reported. The objectives of this study were to: a) evaluate qualitative differences in the uterine flush fluid proteome between mares in oestrus and mares in dioestrus and b) perform a functional classification of proteins either unique to each stage or common between the two stages. Uterine flush fluid samples were collected from 8 light breed mares in either oestrus (n = 5) or dioestrus (n = 3). Proteomic analysis of the samples was conducted using liquid chromatography-tandem mass spectrometry. Proteins exclusively detected in oestrus or dioestrus and those common to both stages were identified using the Scaffold software (version 4.4.8, Proteome Software Inc., Portland, OR). The identified proteins were classified into gene ontology (GO) categories (cellular component [CC], molecular function [MF] and biological process [BP]) using the PANTHER (www.pantherdb.org) classification system version 14.0. Of 172 proteins identified, 51 and 28 were exclusively detected in mares in oestrus and dioestrus, respectively, and 93 proteins were common to both stages. The most represented terms in various GO categories were similar among the three subsets of proteins. The most represented CC terms were extracellular region and cell, the most represented MF terms were catalytic activity and binding, and the most represented BP terms were metabolic process and cellular process. In conclusion, proteomic analysis of the uterine flush fluid enabled the identification of subsets of proteins unique to oestrus or dioestrus, or common to both stages. The results of this study can serve as a baseline for future research focused on finding stage-specific protein markers or evaluating differences in the uterine flush fluid proteome between normal mares and those with uterine disease.

A comparison of the uterine proteome of mares in oestrus and dioestrus.

Proteomic analysis of mare uterine flush fluid provides a minimally invasive technique for studying protein changes associated with the oestrous cycle. The aim of this study was to identify differentially abundant proteins in the uterine flush fluid of mares in oestrus and dioestrus. In this study, uterine flush fluid samples were collected from eight reproductively healthy mares in either oestrus (n = 5) or dioestrus (n = 3). Proteomic analysis was performed using liquid chromatography-tandem mass spectrometry. Of 172 proteins identified, six proteins (immunoglobulin lambda-like polypeptide 1, haemoglobin subunit alpha, alpha-1B-glycoprotein, serotransferrin, apolipoprotein A-1, and haemoglobin subunit beta) were significantly more abundant in oestrus. These proteins may contribute to the endometrial defence system through roles in inflammation, immunity or antimicrobial activity. In other species, some of these proteins have been described as immunoglobulins, negative acute phase proteins or defence agents against micro-organisms. During dioestrus, immunoglobulin alpha-1 chain C region-related, complement factor I, CD 109 antigen and uterocalin, were significantly more abundant. Research in other species suggests that these four proteins contribute to the immune response through proposed immunoregulatory characteristics, complement system involvement or roles in B cell-T cell interactions. In conclusion, ten differentially abundant proteins were identified in the uterine flush fluid of mares in oestrus and dioestrus. Targeted studies on these proteins could elucidate their role in uterine defence mechanisms during the oestrous cycle in the mare.

Singleton pregnancy with concurrent pyometra in the contralateral horn in a bitch with a live puppy outcome.

A 20-month-old, 48-day pregnant female American Bulldog was presented for intermittent bloody vaginal discharge of 4-day duration. The bitch was clinically healthy with a mild neutrophilia. Transabdominal ultrasound revealed a viable foetus in the left uterine horn and fluid in the right uterine horn. She was treated with amoxicillin/clavulanic acid and re-examined at regular intervals to monitor for deterioration of the bitch or foetus. The uterine fluid progressively decreased in volume, and the puppy remained viable. A Caesarean section was performed on the bitch's expected due date, resulting in a healthy puppy. This is only the second published report of a pregnancy and concurrent pyometra with a live puppy outcome. This case is unique in that the bitch's pyometra resolved grossly during pregnancy.

Inflammatory markers for differentiation of endometritis in the mare.

BACKGROUND: Endometrial biopsy is required to diagnose mares with chronic endometritis and endometrial degenerative fibrosis. An increase in understanding of equine reproductive immunology could be utilised to create less-invasive, time-efficient diagnostic tools especially when evaluating mares for chronic endometritis. OBJECTIVES: To evaluate inflammatory cytokine and chemokine concentrations in uterine fluid samples collected by low-volume lavage (LVL) as a potential screening diagnostic biomarker for endometritis. STUDY DESIGN: Prospective cross-sectional clinical study. METHODS: Forty-six mares underwent a LVL and subsequently endometrial biopsy. Mares were split in three groups: healthy, acute endometritis, and chronic endometrial fibrosis (CEF) based on cytological and histological evaluation. A fluorescent bead-based multiplex assay for IFN-γ, IFN-α, IL-1ß, IL-4, IL-10, IL-17, sCD14, TNF-α, CCL2, CCL3, CCL5 and CCL11 were carried out on the LVL fluid. The endometrial biopsy was utilised for histology and qPCR of IFN-γ, IL-1ß, IL-6, IL-8, IL-17, TNF-α, CCL2 and CCL3 genes. Statistical analyses examined differences in inflammatory markers and predictive modelling for diseased endometrium. RESULTS: Secreted concentrations of IFN-γ were lower in LVL fluid from reproductively healthy mares compared with acute endometritis (p = 0.04) and CEF (p = 0.006). Additionally, IL-17, IL-10, IL-1ß, TNF-α, CCL2, CCL3, CCL5 and CCL11 were significantly increased (p ≤ 0.04) in LVL from CEF mares compared with healthy mares. Mares with CCL2 concentrations ≥550 pg/mL (14/14) had 100% probability of having CEF and/or acute endometritis. Healthy mares had lower relative abundance of IL-17 mRNA compared with mares in CEF group [median (interquartile rage) = 14.76 (13.3, 15.3) and 12.4 (10.54, 13.81)], respectively (p = 0.02). MAIN LIMITATIONS: Limited sample size: larger numbers of mares with and without endometritis are required and reference intervals in LVL samples have to be established. CONCLUSIONS: Inflammatory chemokines and cytokines concentrations differed between healthy mares and mares with acute endometritis or CEF in LVL.

Serum progesterone and oxytocinase, and endometrial and luteal gene expression in pregnant, nonpregnant, oxytocin, carbetocin and meclofenamic acid treated mares.

Our objectives were to examine changes in endometrial and luteal gene expression during estrus, diestrus, pregnancy and treatments to induce luteolysis and putatively induce luteostasis. Groups were: Diestrus (DIEST), Estrus (ESTR), Pregnant (PREG), Oxytocin (OXY), Carbetocin (CARB), and Meclofenamic acid (MFA). Blood was obtained from day (D)12 to D15 for measurement of oxytocinase, also referred to as leucyl-cysteinyl aminopeptidase (LNPEP) and progesterone. Luteal biopsies were obtained on D12 and D15 and an endometrial biopsy on D15. Real-time RT-PCR was performed for the following genes: PGR, ESR1, OXTR,OXT, LNPEP, PTGS2, PTGFR, PLA2G2C, PTGES, SLC2A4, and SLC2A1. Regarding serum LNPEP, PREG and OXY (p-value<0.001) had higher concentrations than DIEST mares. Endometrial PTGES expression was higher (p-value <0.04) in DIEST, PREG and OXY than other groups. Endometrium from ESTR had increased expression of OXT (p-value < 0.02) compared to MFA and OXY mares. Carbetocin treatment: decreased serum progesterone and LNPEP; increased endometrial PLA2G2C; decreased endometrial PTGES; and decreased luteal aromatase and PTGES. Treatment with MFA: decreased endometrial PLA2G2C, increased endometrial PTGES; and resulted in less OXTR and OXT luteal abundance on D12 compared to D15. Endometrial and luteal expression of LNPEP is affected by physiologic stage and treatment and is involved in luteal function and pregnancy recognition pathways through effects on oxytocin and prostaglandin synthesis in the horse.

Characterization of serum and tissue oxytocinase and tissue oxytocin in the pregnant and non-pregnant mare.

Oxytocin is a hormone with functions in: reproduction, maternal bonding, milk ejection, and feeding/social behavior, and is reported to be present in a variety of tissues. Our goal is to characterize oxytocin and leucyl and cystinyl aminopeptidase (LNPEP/oxytocinase), a key regulator of oxytocin in mares. We measured serum and tissue LNPEP by ELISA from ovulation (D0) until D21-22 in non-pregnant (n = 5) and pregnant mares (n = 6); and in periparturient and postpartum mares (n = 18). Placenta (n = 7) and homogenized tissue of diestrus mares (n = 6) were evaluated using protein determinations and LNPEP ELISAs. Identification of LNPEP and OXT protein in tissues was also performed via western blot, immunohistochemistry and liquid chromatography-mass spectrometry (LC-MS/MS). Furthermore, in situ hybridization was performed for LNPEP and OXT on endometrium, myometrium, pituitary and corpus luteum (CL). Serum LNPEP concentration were similar. Placental LNPEP U/mg protein was highest in the body and pregnant horn. The highest to lowest LNPEP U/mg protein by tissue were: myometrium > follicle wall > endometrium > kidney > CL > liver. Oxytocin was identified in the equine pituitary, CL and placenta and is likely to act in autocrine or paracrine manner, while LNPEP may act systemically and locally to regulate the availability of OXT.

Rho-associated coiled-coil containing kinase inhibitor improves outcomes of direct-transfer slow-cooled bovine blastocysts.

Direct-transfer slow-cooling cryopreservation is a widely used method for bovine embryo cryopreservation. However, the transfer of cryopreserved embryos is associated with reduced pregnancy rates. Rho-associated coiled-coil containing kinase inhibitor (ROCKi) has shown promise in improving the viability of post-warmed vitrified bovine embryos. Our objective was to investigate the effects of ROCKi treatment prior to slow-cooling or after cryopreservation on embryo viability. In vitro produced bovine embryos (n = 571) were randomly assigned to one of five groups: No-cryopreservation control group (NC-C), C-C group were cryopreserved by slow-rate cooling without ROCKi at any point, R-C group were incubated with ROCKi for 2 h before cryopreservation, C-R group were not exposed to ROCKi prior to cryopreservation but were cultured with ROCKi after cryopreservation, and R-R group were exposed to ROCKi before and after cryopreservation. Treatment group was significantly associated with blastocoel re-expansion, hatching, and degeneration (P < 0.0001). Blastocoel re-expansion rates were lower (P < 0.05) in the C-C (75.2 ± 4.2%) and R-C (85.2 ± 4.7%) groups compared with the NC-C (99.0 ± 0.7%), C-R (94.7 ± 2.6%) and the R-R (94.5 ± 2.9%) groups. The median time to re-expansion was significantly slowest in the C-C group (650, 560-915 min), followed by the R-C group (538, 421-611 min), then the C-R and R-R groups were similar (291, 261-361 and 321, 271-371 min) and the NC-C group was the fastest (196, 161-230 min) (P < 0.05). Similarly, the post-thaw hatching rate was lower, and the median time to hatching slower in the C-C (58.1 ± 7.0%, 2,033, 1634-2820 min) and R-C (65.7 ± 6.9%, 1,853, 1494-2356 min) groups compared with the NC-C (81.7 ± 6.0%, 1,309, 1084-1514 min), C-R (77.2 ± 6.5%, 1,384, 1013-1754 min) and R-R (82.0 ± 5.3%, 1,209, 943-1424 min) groups. ROCKi supplementation after cryopreservation resulted in fewer degenerated embryos (C-R = 8.9 ± 2.8%, and R-R 7.1 ± 2.8%) compared to the C-C (26.8 ± 4.3%) and R-C (17.9 ± 5.7%) groups. Exposure to ROCKi both before cryopreservation and after-cryopreservation yielded the best outcomes, similar to NC-C control group without cryopreservation, and significantly better than the C-C control group without supplements. Exposure to ROCKi after cryopreservation demonstrated greater benefits compared to exposure before cryopreservation alone. These findings suggest that ROCKi can potentially enhance cryosurvival of bovine embryos.

Expression of Oxytocin/Neurophysin I and Oxytocinase in the Equine Conceptus from Day 8 to Day 21 Post-Ovulation.

Leucyl and cystinyl aminopeptidase (LNPEP/oxytocinase) is an enzyme that metabolizes oxytocin in serum and tissues. The presence of oxytocin/neurophysin I (OXT), oxytocin and LNPEP and their relationship to other genes is unknown in the equine conceptus. Our objective was to characterize gene expression of LNPEP and OXT on D8, 10, 12, 14, 15, 16 and 21 conceptuses in relationship to other genes. Immunohistochemistry, western blot and liquid chromatography with tandem mass spectrometry (LC-MS/MS) were used for identification of oxytocin and LNPEP in D15, 16 and 18 conceptuses. LNPEP was increased at D15 compared to D10, was immunolocalized in the equine trophectoderm and endoderm, and protein was confirmed by LC-MS/MS. Maximal abundance of OXT was at D21, and lowest on D12 and D14, but no protein was identified. OXTR abundance was highest on D14 and D21. LNPEP was correlated with PTGFR and PTGES on D12 and D14-D15, and high expression of PTGES, PTGS2 was found on D14, D15 and D21; PTGFR was found on D8 and D12-21. LNPEP may have a role in prostaglandin regulation and conceptus fixation by decreasing the availability of oxytocin. Further investigation on the role embryonic LNPEP during pregnancy is warranted.

Endometrial and luteal gene expression of putative gene regulators of the equine maternal recognition of pregnancy.

Our understanding of the temporal changes in endometrial and luteal gene transcripts related to the actions of oxytocin and prostaglandin during early equine pregnancy is incomplete. Additionally, the role of oxytocinase, also known as Leucyl-cystinyl aminopeptidase (LNPEP), during early pregnancy in mares has not been previously investigated. Luteal and endometrial biopsies were obtained on Day (D)8, D10, D12 and D15 post-ovulation in pregnant (PREG) and diestrus (DIEST) mares for real-time qPCR. Differences in endometrial gene expression occurred over time in: SLC2A4, SLC2A1, PTGES, OXTR and LNPEP. PTGFR and PLA2G2C had lower relative abundance in PREG D15 endometrium compared to D10. OXT and OXTR were increased on D10 and 15 PREG, respectively. Regarding luteal mRNA relative abundance, ESR1, PTGS2, PTGFR, and PTGES had higher relative abundance in D12 of DIEST and PREG. Luteal expression of OXTR and OXT had higher relative abundance in D15 compared to D8, and LNPEP had higher relative abundance in D10 and 12. Endometrial and luteal PTGES had an increased mRNA abundance in both D12 DIEST and PREG mares, which may lead to additional luteoprotective prostaglandin E2 (PGE2) secretion. Furthermore, luteal SLC2A1 had higher relative abundance in pregnancy, and likely supports the high metabolic activity of luteal tissue by increasing glucose uptake. Oxytocinase is present in endometrial and luteal tissue and its role in oxytocin induced prostaglandin secretion is uncertain.

Malunion of an In Utero Metacarpal Fracture in a Standardbred Mare Presenting for Dystocia.

In utero fracture and malunion of long bones is a rare condition in horses. Most foals with in utero fractures are aborted, and the identification of a fetal in utero fracture in a mare with dystocia has not been reported. A 7-year-old multiparous Standardbred mare presented to a referral center for correction of dystocia. Assisted vaginal delivery and controlled vaginal delivery attempts were unsuccessful mainly because of contracted tendons impeding mutation. As the foal was alive, a cesarean section was elected. The foal was delivered but ultimately euthanized because of the congenital abnormalities. Computed tomography of the right forelimb of the foal along with gross examination and histologic evaluation of the right metacarpus revealed the malunion of a previous in utero fracture. While a few cases have been reported of in utero fracture, many of these were in abortuses and not in fetuses at term, making this case a new presentation and potential etiology for dystocia.

Incidence and risk factors for canine mastitis and metritis in two guide dog populations.

Canine mastitis and metritis can cause severe illness but the incidence and risk factors have not been well-studied. Goals in the present study were: 1) report the incidence of mastitis and metritis in a large population, and 2) identify potential risk factors that predispose females to those diseases. A retrospective cohort study was conducted using data from two guide dog colonies that was collected for 17 and 10 years, respectively, for the two colonies. A total of 3076 whelpings occurred during the respective study periods and data were analyzed. Clinical mastitis was diagnosed in 13.2 % of whelpings (408 cases) with the average day of diagnosis being 16.7 postpartum. Risk factors for mastitis identified were colony, litter size where bitches that had large litter size of ≥9 pups (16.2 %) were 60 % more likely to develop mastitis compared with bitches that had litters of <9 pups (11.3 %). Bitches with congestion of the mammary gland were 4.8 times more likely to develop mastitis compared with bitches without mammary congestion. Case incidence of metritis was small (0.7 % of whelpings) and occurred on average at day-5 postpartum (range 1-16). There were no significant risk factors identified, and this may be due to the small number of metritis cases (22 cases) in the present study. Interpretations regarding metritis, therefore, should be made with caution. The results from this study provide parameters for breeders and veterinarians to identify bitches that may require close monitoring for mastitis and metritis.

Impacts of oocyte/zygote timing for in vitro fertilization and gene editing in the dog.

Previously, we reported the first live births of dogs using in vitro fertilization (IVF), embryo cryopreservation, and transfer. These techniques have potential applications in the conservation of endangered canids, and development of gene editing/repair technologies that could improve animal welfare by restoring normal gene function and removing predisposition to disease. Here, we used IVF as a springboard for initial attempts at genetic modification through gene editing/repair using the Clustered Regularly-Interspaced Short Palindromic Repeat (CRISPR)-CRISPR-associated endonuclease (Cas9) system. We showed previously that timing is critical for successful IVF in that the canine oocyte must be exposed to the oviductal environment beyond simply reaching metaphase II. Others have shown that timing of injection of CRISPR-Cas9 constructs is critical in gene editing, influencing the extent of genetic mosaicism. Therefore, we investigated whether timing of injection of the gene editing/repair constructs might influence the success of embryo production and gene editing in the dog. We achieved similar IVF success to our prior report in generating 2-cell control embryos, and found equally reduced embryo production whether injection was performed in oocytes prior to fertilization, or in presumptive single-cell zygotes already exposed to sperm. We had no success at generating offspring with precise single-nucleotide changes in KRT71 via homology-directed repair (HDR), but did identify mutation of FGF5 using non-homologous end joining (NHEJ). These findings underscore the difficulties inherent to gene repair, but represent important progress on reproducibility of canine IVF, improved techniques of oocyte/embryo handling, and impact of timing of injections on embryo development.

Impact of decision to delivery time of fetal mortality in canine caesarean section in a referral population.

In human medicine, there is a recommended decision to delivery interval (DDI), which allows for the optimization of protocols and systematic review of hospital success. In veterinary medicine, no such guideline has been established or investigated. The purpose of this study was to investigate the relationship between the interval from the decision to perform a caesarean section and the delivery of the neonates and fetal mortality at the time of surgery. One hundred and fifty canine caesarean sections were evaluated retrospectively. Caesarean cases were dichotomized to those that had at least one perinatal death and cases where all puppies survived. Factors that increased the likelihood of at least one perinatal death at caesarean section were: cases that presented as an emergency caesarean section, the dam presenting with a fetus in the vaginal canal, the dam not having a history of previous caesareans sections, and being multiparous. Even though there was no association of DDI with likelihood of having perinatal death, timing related factors that increased the likelihood of having at least one perinatal death at caesarean section were: cases where total anaesthesia time was longer than 2 h; time from induction to start of surgery was longer than 45 min and surgical time longer than 75 min. In conclusion, time is a factor in the success of canine caesarean sections and further research is needed to better define the optimal decision to delivery time in canine caesarean sections in order to optimize fetal survival and hospital protocol success rates.

Comparison of Foaling Prediction Technologies in Periparturient Standardbred Mares.

The ability to predict impending foaling is invaluable, as attending foaling improves mare and foal outcome during parturition; therefore, a comparison of foaling prediction technologies is warranted. The purpose of this study was to compare technologies to predict foaling. Forty Standardbred mares were monitored daily for parturition. Prefoaling mammary secretion was collected daily and evaluated using a: calcium titration method (Ca titration), calcium and magnesium strip (Ca and Mg), handheld digital pH device (pH digital), pH strip, and refractometer (Brix). In addition, a birth monitoring system that was attached in the mare's vulva was studied and all positive alarms (foaling) and negative alarms (false alerts) were recorded. Analysis of variance, correlation, and logistic analysis were used to analyze the different technologies and their associations between parity and foal gender. Receiver operator characteristic analysis for cutoff value and subsequently sensitivity, specificity, positive, and negative predictive values for each technology were obtained. Maiden mares and mares carrying a colt had a longer gestational length. Multiparous and maiden mares had no difference in their prefoaling mammary secretion pH using the digital and pH strip technologies when predicting foaling; however, the calcium concentrations measured using the Ca titration technology were lower in multiparous than in maiden mares. The Ca titration technology was the single best technology to predict when the mare was not going to foal; and when the Ca titration and pH digital were used in combination, their prediction for foaling increased when compared with using each technology individually.

The Effect of Routine Postpartum Uterine Lavage on Endometrial Cytology, Culture, and Pregnancy Rates in Thoroughbred Broodmares.

Minimizing the interval between parturition and successful rebreeding of thoroughbred broodmares is important in maintaining annual foal production and broodmare profitability. Routine postpartum uterine lavage may help achieve this by aiding uterine clearance and involution in the early postpartum period and increasing pregnancy rates. This retrospective study investigates the effect of postpartum uterine lavage in 32 thoroughbred mares housed on a single farm in Australia in 2014 and 2015. In the control year (2014), postpartum uterine lavage was not performed. In the treatment year (2015), all mares received a uterine lavage 3 days postpartum. In both years, endometrial swabs were obtained for culture and cytology at the first postpartum estrus (foal heat swab) and the second postpartum estrus (second heat swab), prior to the mare being bred by natural cover at the second postpartum estrus. There was no significant difference in bacterial culture rates from foal heat or second heat swabs between the control and treatment years. Assessment of endometrial cytology of the foal heat swab and second heat swab revealed a significantly higher proportion of polymorphonuclear cells (PMNs) in the control year compared with the treatment year. Positive culture and cytology rates were significantly higher at foal heat than at the second postpartum heat in both years. No association of year, cytology, or culture with pregnancy rates was identified. In conclusion, routine postpartum uterine lavage significantly reduces the presence of endometrial PMNs at first and second postpartum estrus but does not reduce endometrial culture rates or increase pregnancy rates.

Efficacy and outcome of foaling augmented with oxytocin using mammary calcium and pH criteria to guide the timing of augmentation.

Augmentation of parturition can be used to advance labor in mares to occur at a time when personnel is available to assist if necessary. We performed a retrospective study to determine the efficacy and safety of augmentation to manage foalings. Augmentation was performed with 3 IU oxytocin i.v. when mammary calcium concentrations were ≥250 ppm, mammary secretion pH ≤ 6.5, and the mare showed impending signs of parturition. Augmented parturitions (n = 19) were compared with three different control groups. The three control groups were: 1) Time Match control (n = 37) which were non-augmented foalings in the barn during the same time period; 2) Mare Match control (n = 32) which were the non-augmented parturitions of the augmented mares in previous years; and 3) Historic Match control (n = 165) consisted of foalings that occurred from 2006 to 2016 in the facility. All augmented mares foaled within two h with an average of 44 min (range 20-75) after oxytocin injection. The interval between foaling and the foal standing was shorter in augmented parturitions compared with historic match controls. The interval between foaling and the foal nursing was longer with augmented parturitions compared with time match and historic match controls. Duration of fetal membrane retention was not different between all groups. Augmentation of imminent parturition is potentially a safe and effective treatment for mares and foals. Implementation of augmentation as a routine procedure may increase the likelihood of enteral administration of colostrum to foals.

Treatment of Hydropsical Conditions Using Transcervical Gradual Fetal Fluid Drainage in Mares With or Without Concurrent Abdominal Wall Disease.

Hydropsical conditions are exceedingly rare in the horse. However, when they occur, they are true emergencies due to the severe enlargement of the pregnant uterus, which can result in clinical signs, such as an enlarged round abdomen, dyspnea, reluctance to walk, and colic, and may lead to the development of abdominal wall disease. The pathogenesis of hydropsical conditions is not fully elucidated, but they have been associated with placentitis and fetal abnormalities. This report describes six cases of hydropsical conditions in mares with or without concurrent abdominal wall disease. Five out of six cases were hydrallantois, and of these five, two mares had abdominal wall disease; the remaining one out of six cases was hydramnios. All mares were treated by termination of the pregnancy through gradual fluid drainage transcervically over a number of hours, and their fetuses were delivered vaginally. All fetuses were euthanized immediately after vaginal delivery. Of the six mares, two had signs of placentitis, two were confirmed seropositive for leptospirosis, and two were euthanized (one because of a vaginal tear that communicated through the peritoneum and one mare that developed abdominal wall rupture and laminitis). The remaining 4 mares were available for follow-up; three mares were not rebred, and one mare became an embryo donor, with a successful embryo recovery. We reported the prevalence of leptospira involvement in two out of six cases of hydrallantois and also described the clinical outcome of the mares after treatment with slow fetal fluid drainage.

Progesterone levels and days to luteolysis in mares treated with intrauterine fractionated coconut oil.

Intrauterine plant oil infusion, including fractionated coconut oil, has been previously found to be a safe, inexpensive, and reversible method of prolonging the luteal phase in mares when administered on Day 10 of the estrous cycle. Our objective was to understand the uteroovarian response to the administration of fractionated coconut oil infusion in the uterus of diestrous mares. We hypothesized that intrauterine coconut oil administration on Day 10 would prolong luteal life span in a dose-dependent fashion and would result in higher serum progesterone levels than untreated mares at the expected time of luteolysis. Light-horse mares (n = 18) were examined using transrectal palpation and ultrasonography to determine if they had a normal interovulatory interval and were then examined daily in estrus until the day of ovulation (Day 0) and then every other day during an estrous cycle. Jugular blood was drawn on Day 11, Day 13, Day 15, and Day 17, centrifuged, and serum stored until assayed for progesterone (P4; Siemens Coat-a-Count Progesterone RIA, Los Angeles, CA, USA). Mares were randomly assigned to treatment and studied over one to two estrous cycles with a rest cycle after each treatment cycle. Groups were: control (n = 5), fractionated coconut oil 1.0 mL (Miglyol 810; Sasol Oil, Witten, Germany) infused in the uterus with an artificial insemination pipette on Day 10 (Group 1; n = 5) and fractionated coconut oil 0.5 mL infused in the uterus with an embryo transfer gun, on Day 10 (Group 2; n = 5). All statistical analyses were performed using analytical software (Stata SE, version 13.1, College Station, TX, USA) at P < 0.05. Data were first evaluated using the Shapiro-Wilk test for normality. Differences between groups in days to luteolysis (DTL) were examined using analysis of variance and Bonferroni, and the effect of day and treatment on P4 levels were examined using the Kruskal-Wallis and Dunn's all pairwise test. There was a significant difference in DTL between the groups (P = 0.0083), with fewer DTL in Group 1 compared to control (P = 0.011) and to Group 2 (P = 0.034). There was a significant effect of day (P < 0.0001) on P4 levels with Day 11 P4 levels higher than Day 15 and Day 17 and Day 13 P4 levels higher than Day 15 or Day 17 (P < 0.03). There was a significant effect of treatment group (P = 0.0098) on P4 levels with control and Group 2 levels higher than Group 1 mares (P = 0.0012, P = 0.0495, respectively). We concluded that intrauterine administration of 1 mL of fractionated coconut oil lowered P4 levels in diestrus in a dose-dependent fashion and did not prolonged the luteal phase of the mares.

Evaluation of diagnostic utility, safety considerations, and effect on fertility of transvaginal ultrasound-guided ovarian biopsy in mares.

Ultrasound-guided biopsies of corpora lutea have been previously used for research purposes in the mare and cow. However, the health effects and fertility after transvaginal luteal biopsies (TVLB) or transvaginal ovarian biopsies (TVOB) obtained for diagnostic purposes in cases of suspected ovarian tumors have not been previously evaluated in the horse. The aim of this study was to determine the effects on health and fertility of TVLB and TVOB in mares; 53 mares were included in the study (11 control non-biopsied mares, 37 TVLB mares biopsied on one or more of the following Days 8, 10, 12, 15, 21, and 5 TVOB mares with ovarian abnormalities), resulting in a total of 108 TVLB and TVOB cycles and 183 procedures. Mares were divided into Groups 1 to 3 by the number of TVLB per estrous cycle (mare in Group 1 had 1 TVLB procedure, mares in Group 2 had 2 TVLB procedures, and mares in Group 3 had 3 TVLB procedures). Group 4 comprised TVOB mare cycles with ovarian abnormalities (n = 5). Mares were examined to determine day of ovulation (Day 0) and the presence of a corpora lutea using transrectal ultrasonography. Mares were sedated, and an ultrasound-guided transvaginal biopsy was performed of luteal or ovarian tissue. Health effects of TVLB or TVOB were assessed by daily rectal temperatures, appetite, and general demeanor for 72 hours post-procedure, and 3 mares were examined at necropsy. Fertility was not significantly different in control and TVLB Groups 1 to 3 (P = 0.7648) and in the first or subsequent cycles where the ovulation was from an ovary that had a previous TVLB (P = 0.7147). A TVLB on Day 8 post-ovulation may induce an early return to estrus. In conclusion, the TVLB or TVOB procedure had no effect on health and fertility in this study if the procedure was correctly performed with good technique. Because of the low number of cycles (n = 37), the fertility data should be interpreted with caution. The TVOB may be applied in the diagnosis of mares with ovarian abnormalities, and no adverse health effects were associated with TVOB of mares with granulosa theca cell tumor.