Embryonic Vesicle Research Articles

A novel method for equine embryo transfer from contaminated recipient mares into second healthy recipients for surviving embryos

Embryo transfer (ET) is regarded as one of the reproductive methods for genetic improvement initiatives in the horse business. The current study aimed to increase the success rate of ET by re-flushing and re-transferring of embryos from contaminated recipient mares into second healthy recipients. The study involved 25 recipient mares that demonstrated embryonic vesicles inside intraluminal fluid (ILF) on ultrasound scanning 2 ± 0.5 days after ET. Seven recipient mares were left without re-flushing and re-transferring of embryos (control group) and 18 recipient mares (treated group) were subjected to re-flushing and re-transferring of the embryos to second healthy and prepared recipients. The retrieved fluid from treated mares was subjected to microbiological and cytological examinations. The results revealed mixed and single infections, as well as polymorphonuclear neutrophils (PMNs). The total bacterial count and cytology findings revealed that all first recipient mares had endometritis. All control mares showed embryonic death at 14 ± 1.1 days (13–15 days) of embryonic age. In treated group, microscopic evaluation of retrieved embryos revealed a well-developed thick capsule in 12/18 (66.7 %) of them and a normal capsule in 6/18 (33.3 %). The pregnancy rate in well-developed capsule embryos was greater (66.7 %) than in typical capsule embryos (33.3 %). This study illustrates the value of swabbing the uterus of the recipient mares during estrus prior to ET and their checking at 2 days after ET. In addition to boosting pregnancy rates in ET commercial programs, our novel method can protect the embryos from the harmful effects of the inflammatory process and help them survive.

Development of the first equine blastocyst produced by conventional IVF and in vitro culture in Europe resulting in the birth of a foal

In this case report, the production of the first equine blastocyst using conventional in vitro fertilization (IVF) and in vitro embryo culture methods in Europe is described. A healthy foal was born after transfer of a blastocyst using a complete in vitro production process, including in vitro oocyte maturation, in vitro sperm capacitation, fertilization in vitro and culture to the blastocyst stage in vitro. Oocytes were recovered from ovaries of slaughtered mares. After in vitro maturation, the oocytes were fertilized by conventional IVF and cultured for nine days. One embryo reached the blastocyst stage and was vitrified. After the selection of a suitable recipient mare that had ovulated four days earlier, the blastocyst was thawed and transferred. Five days after embryo transfer, a single embryonic vesicle was detected by transrectal ultrasonography. After a normal pregnancy of 323 days, a healthy colt was born. Parentage testing via microsatellite genotyping confirmed that the recipient was excluded as the foal’s dam and that the stallion whose semen was used, qualified as the sire.

Transvaginal aspiration, oocyte maturation and production of blastocysts and a pregnancy via ICSI in an endangered breed: Baudet du Poitou donkey

Two Poitou donkey jennies were presented for clinical oocyte recovery and embryo production via intracytoplasmic sperm injection (ICSI). Both jennies underwent transvaginal ultrasound-guided follicle aspiration on two occasions. Recovered oocytes were held overnight then placed into maturation culture, using standard methods for mare oocytes. On the first replicate for both jennies, the oocytes were divided into two groups; one group was denuded and examined at 30 h culture (standard culture duration for mare oocytes) and the second was denuded and examined at 36 h culture. No oocytes with polar bodies were observed at either time. The oocytes were maintained in maturation culture until 46 h, at which time oocytes with polar bodies were observed. Semen was then prepared; oocytes underwent ICSI approximately 48 h after being placed into maturation culture. On the second replicate for both jennies, oocytes were cultured for maturation for 42 h, then denuded and subjected to ICSI at 46 h. Sperm preparation, injection and embryo culture were performed as for mare oocytes. Blastocyst rates per injected oocyte were 8/19 (42 %) overall, being 4/12 and 4/7 for the first and second TVAs, respectively. Blastocysts were vitrified. Three blastocysts were warmed and transferred to Poitou donkey jenny recipients. One embryonic vesicle was visualized on ultrasonography on embryo Day 12, which increased in size on Day 13 but was not present when examined on Day 14. These results demonstrate that oocyte recovery and ICSI are efficient for production of Poitou donkey blastocysts. To the best of our knowledge, this is the first report of production of blastocysts via ICSI in the Poitou donkey, and the first report of transfer of ICSI-produced embryos in the donkey. Further work is needed on factors affecting pregnancy after embryo transfer in the donkey.

The effectiveness of Ultrasonography in predicting Pregnancy, litter size, viability and embryo/fetal age during first three months post-breeding in Iraqi Shami does

This study aimed to evaluate the detection efficiency of early pregnancy by Ultrasonography (ULR), and also to estimate embryo/fetal age, embryo/fetal count and viability in Iraqi Shami Does. Thirty-five mature does were synchronized using progesterone intra-vaginal sponges for 12 days+ equine Chorionic Gonadotrophin (eCG) (500 IU) after sponge removal. Estrus was detected after two days in only 28 does, therefore, seven does were excluded. All does (n=28) were mated by four breeding bucks. Pregnancy monitoring, embryo/fetus number and developmental stages during the does gestation was carried out with B-mode Transrectal ultrasonography (TRU) (Transrectal probe) on days 17-29 Post mating (PM) using a frequency 6.5-7.5 MHz, and Transabdominal Ultrasonography (TAU) with in days 36-90 PM with frequency 3.5-4.5 MHz. The results revealed that the conception rate was 85.71% (24/28) and the Predicted +value (PV+), Predicted –value (PV-), Sensitivity (Se), Specificity (Sp) and Accuracy (Acc) were low on days 17-20 PM, which recorded 50%, 4%, 12%, 25% and 14% respectively. All detection values between 96-100% in the period 36-50 PM, and 100% in day 60 onwards. The efficiency in predicting embryo/fetal number was 77% and 75% for estimating twin (Se) and singleton (Sp) pregnancies in days 23-29 PM, and the detection parameters reached 100% by day 60, then declined after that. The embryo/fetal viability was 60% in the early gestational period and reached 96% between days 36–50 PM, and 100% after that. The correlation coefficient between embryo/fetal age and Embryonic Vesicle Diameter (EVD), Crown Rump Length (CRL) and Trunk Diameter (TD) was recorded as very high in different gestational ages; R² = 0.99 (23-42 days), R² = 0.97 (23-60 days) and R² = 0.98 (42-90 days). It was concluded that ULR (by using both TRU and TAU) was reliable, very efficient and safe to predict pregnancy, embryo/fetal count, viability and age between days 23-35 and 35-90 PM, respectively.

Power-doppler ultrasonography is an accurate tool for early pregnancy detection in donkeys

Early detection of pregnancy could enhance the reproductive efficiency of donkeys, as the species has a wide range for the first detection of the embryonic vesicle (EV) via B-mode ultrasonography. This study aimed to assess the uterine blood flow of jennies as an early indirect pregnancy diagnostic test. We hypothesized that uterine blood flow is increased, thus, serving as an early pregnancy diagnostic tool. Jennies (n=52) were bred with fresh semen from a fertile jack, and nine jennies were not bred and served as controls; ovulation (Day-0) was confirmed at 12h intervals for all jennies. Power-doppler and B-mode ultrasonography were performed on Days 7 to 10 to assess the uterine horn blood flow and identification of EV, respectively; the latter was repeated on Day 15. Uterine horn blood flow was evaluated using spot meter techniques, measuring mean pixel intensity. Progesterone concentrations were determined with RIA on Days 7 to 10. Data were assessed for normality with Shapiro- Wilk's test and mixed models. Receiving operating curve was used to determine the cut off value for uterine blood flow. Sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) for blood flow on Days 7 to 10 were calculated to identify pregnant jennies and confirmed on Day 15. Significance was set as P≤0.05 and tendency as 0.05<P<0.1. Of 52 bred jennies, 29 (53%) were confirmed pregnant on Day 15. B-modeultrasonography identified 17.2% (5/29) of the EVs on Day 9 and 48.3% (14/29) on Day 10; no EVs were identified on Days 7 to 8. Pregnant jennies (n=29) had greater uterine blood flow in comparison with non-pregnant jennies (144.8±1.5 vs. 139.2±1.3 pixels/unit, n=32) (P=0.007), with an SD of the difference of 6.3 pixels/unit (p=0.008). There was no effect of time (P=0.48) or group x time interaction between (P=0.70) for uterine blood flow. Progesterone concentrations did not change within Days 7 to 10 (P=0.81), and there was no group x time interaction (P=0.49). Progesterone tended to be greater in pregnant than non-pregnant jennies (P=0.07). There was no effect of time (P=0.79), group (P=0.77), or group x time interaction (P=0.92) for uterine horns. Sensitivity from Days-7-to-10 were 86%, 82%, 60%, and 62%, respectively. In contrast, specificity for Days 7 to 10 was 80%, 88%, 78%, and 75%, respectively. The PPV and NPV for Days 7 to 10 were 81% and 85%; 75% and 20%; 35% and 33%; and 30% and 27%; respectively. In conclusion, this study demonstrated the efficacy of power-doppler-ultrasonography as a sensitive and specific tool for early indirect detection of pregnancy in donkeys, whereas standard B-mode first detected a small percentage of pregnancies on Days 9 to 10. Power-doppler could facilitate the breeding management of donkeys, such as timing the induction of luteolysis in non-pregnant jennies at Days 7 to 10 post-ovulation.

Successful mule embryo transfer to a recipient mule during late diestrus

Mating between a donkey (Equus asinus) and a mare (Equus caballus) results in a mule foal, which is an infertile hybrid due to the different number of chromosomes in the parent species. However, mules have been successfully used as recipients of donkey embryos, horse embryos and, recently, mule embryos. In non-cyclic recipient mules, progesterone supplementation is needed to sustain pregnancy, but some mules can develop pre-ovulatory follicles which ovulate and form active CLs that produce enough progesterone to sustain pregnancy. In spite of this, it is possible that the induction of additional CL formation to increase progesterone concentrations prior to embryo transfer could increase the chances of successful pregnancy. In this study an 8-day old mule embryo produced by the insemination of a mare with fresh donkey semen was transferred to a cyclic mule that had been induced to form a secondary CL by the injection of 1.8mg of deslorelin acetate on day 8 post-ovulation, when a spontaneously developed follicle reached 38 mm of diameter. Progesterone concentrations at that time confirmed that the primary CL was active. Secondary ovulation occurred 2 days later and a secondary CLformed during the following days, resulting in a significant increase in blood progesterone concentrations. The 8-day old embryo was transferred nine days after the secondary ovulation (19 days after the primary ovulation), when both the primary and secondary CL were fully active as indicated by sustained progesterone concentrations and by their diameters of 37 and 35.5 mm, respectively. Pregnancy diagnosis was made 4 days after transfer of the embryo, that is, 23 days after the primary ovulation and 13 days after the secondary ovulation. At that time, the 12-day old embryo was within a 15mm embryonic vesicle. On days 55 and 58 of pregnancy two new supplementary CL formed spontaneously, reaching diameters of 36 and 29mm. This report of a successful mule-in-mule pregnancy using as a recipient a mule with both a primary and an artificially induced secondary CL suggests the possibility of improving the chances of successful pregnancy in recipient mules by increasing endogenous progesterone production through induction of secondary ovulations prior to embryo transfer. Acknowledgements. This study was supported by grant PAPIIT IN 223920, DGAPA-UNAM.

63 Impact of Supplementing Rumen-Protected Methionine During the Periconceptional Period in Fetal and Postnatal Growth in Beef Cattle

Abstract Previous evidence has suggested that fertilization and preimplantation embryonic development are sensitive periods that can be influenced by changes in maternal nutrition. We hypothesized that feeding rumen-protected methionine (RP-Met) during the periconceptional period will program bovine gestation in a manner that enhances fetal and postnatal growth. A total of 114 cows were fed a roughage-based diet and randomized to receive corn gluten supplemented with 15 g of RP-Met (RP-Met; Smartamine, Adisseo) or not (Con) from d -7 to +7 relative to artificial insemination (AI) with sex-sorted semen. Estrus synchronization was conducted using the 7-day CoSynch + CIDR protocol. Pregnancy diagnosis, embryo, and fetal morphology were conducted using transrectal ultrasonography 30, and 60 days after AI. After calving, 40 calves (Cont = 19; RP-Met = 21) were considered for birth weight (BW) analysis. Female calves (n=34) were weighed at 2 months of age. Pregnancy per AI at day 30 (Con = 50%, RP-Met = 55%) and day 60 (Con = 50%; RP-Met = 53.4%) were not different between groups (P ≥ 0.05). At 30 days of pregnancy, embryonic vesicle diameter (Con=11.0 ± 0.7 mm; RP-Met =11.6 ± 0.7 mm; P &amp;lt; 0.05) and circumference (Con=0.88 ± 0.09 cm2; RP-Met =0.99 ± 0.09 cm2 P &amp;lt; 0.05) were larger in the RP-Met group but embryo length and abdominal cavity were not different between groups. No difference was observed in fetal morphology at d 60. A treatment by sex interaction was observed (P =0.04). Male calves (n=6) from cows fed RP-Met showed greater BW than those from Con group (Con=31.9 ± 2.3 kg; RP-Met =41.4 ± 2.3 kg) but no difference was observed in females (n=34). There was no difference in BW at 2 months of age between groups. In conclusion, using RP-Met increased embryonic vesicle diameter and circumference, and BW in males.

Effect of peri-conception high-fat diets on maternal ovarian function, foetal and placentome growth, and vascular umbilical development in goats.

The objective of this study was to determine whether a high-fat diet (HFD) fed to goats for a brief period during peri-conception would optimize reproductive and foetal responses. Thirty-four Anglo-Nubian crossbred adult goats were allocated into three groups: control (n=11), fed with a total mixed ration (TMR) based on chopped elephant grass and concentrate; HFBM (n=11), given TMR supplemented with soybean oil on a 0.5% dry matter basis for 11 days starting nine days before mating (BM); and HFAM (n=12), fed with soybean oil included in the TMR for 15 days after mating (AM). The TMR diets differed in their fat content (7.5% vs. 2.9%). All goats had oestrus synchronized for 14 days BM by intravaginal administration of 60 mg MPA sponge for 12 days. Forty-eight hours BM, the sponge was removed and 0.075 mg PGF2α was applied intramuscularly. After 36h, 1ml GnRH was administered intramuscularly, and goats were mated after sponge removal. The fat groups showed lower feed intake (p < .001) and higher cholesterol levels (p < .001) when HFD was administered. Doppler and B-mode ultrasound evaluations revealed a greater (p < .05) number of small (<3 mm, 10 ± 0.6 vs. 8 ± 0.5) and large (≥3 mm, 6 ± 0.4 vs. 5.0 ± 0.2) follicles and intraovarian blood area (p < .05) in the HFBM group during sponge removal (57.6%) and mating (24.2%) than those of the no-fat group. During AM, the fat-fed groups exhibited higher glutathione peroxidase levels (p < .05) and a reduction (p < .001) in corpus luteum size (19%) and vascularized Doppler area (41%). No difference (p > .05) between groups was found in foetal traits, placentome and umbilical vascular development, except for the embryonic vesicle where HFAM twin pregnancy showed a smaller size than the control (26.1 ± 3.5cm vs. 33.7 ± 2.7cm; p < .01). Thus, HFD applied during peri-conception of goats has no impact on later foetal development but improved the follicular growth when given before the mating. Thus, the use of HFD in periconception has no impact on foetal development but increases follicular growth before breeding time.

Reproductive efficiency of sows inseminated at single dose fixed time with refrigerated, cryopreserved and encapsulated spermatozoa.

The aim was to assess the reproductive efficiency of different techniques used to preserve spermatozoa in artificial insemination semen doses (AI-doses) by evaluating refrigeration at 15°C, cryopreservation and encapsulation. Forty-two hyperprolific sows were treated with buserelin and inseminated once at a single fixed time. The fertility rate, embryonic vesicles viability and the early embryonic mortality (arrested conceptuses) evaluated post-mortem at 24th day of pregnancy, were analysed in order to assess the effectiveness of each proposed technique. Results show an overall reduction on fertility using the three proposal sperm preservation techniques (69.27%, 60.00% and 78.75% for refrigerated, frozen-thawed and encapsulated AI-doses, respectively). Total number of embryonic vesicles was very similar among the three treatments; yet, the number of viable vesicles was numerically different among groups, and thus, embryonic viability was 79.25%, 80.0% and 87.15% for refrigerated, frozen-thawed and encapsulated AI-doses, respectively.

Comprehensive study on dynamics of early embryonic development in Marwari mares

Horse is a seasonal breeder with reproductive activity being associated with long days, i.e. occurring in spring and early summer. Early equine pregnancy shares many features with that of more intensively assessed domestic animals species, but there are also characteristic differences. In mares, early diagnosis of pregnancy can be done due to the spherical shape. The ultrasonic visualization of conceptus is possible from day 9 of equine pregnancy. In spite of numerous, substantial advances in equine reproduction, many stages of embryonic and foetal morphological development are poorly understood, with no apparent single source of comprehensive information. Hence, in the present study we aimed at studying the dynamics of early embryonic developments in equines, particularly for Marwari breed of horses. The results from the current study revealed that, embryonic vesicles (EV) were first observed on 11th day, fixation occurred after 16.25±0.25 days of ovulation, EV lost spherical shape on 15.83±0.38 days, became oval on 17–18th days and irregular on 19–20th days. Heartbeat, detection of allantoic sac, start of foetal ascent, start of foetal descend and end of foetal descend were detected on 23.91±0.41, 23.58±0.35, 38.66±0.28, 40.66±0.33 and 47.33±0.28 days, respectively. In conclusion, early detection of embryonic vesicles and continuous changes observed in conceptus structure indicate positive pregnancy, viability of embryo and are much beneficial in preventing economical loss of equine owner and stable farm.

Ultrasonographic and macroscopic study of pregnancy in golden hamster

BackgroundHamster is widely used as an experimental model in the study of reproductive system. However, pregnancy diagnosis and aging always have been a challenge. ultrasonography have been used in diagnosis of pregnancy in some small laboratory animals, such as rabbits, rats, and mice. Current study describes use of trans-abdominal ultrasonography for pregnancy diagnosis and fetal age estimation in golden hamster. Furthermore, a macroscopic examination was performed to evaluate the embryonic vesicle diameter, crown-rump length, and fetal head diameter. Ten adult female golden hamsters were selected and maintained under controlled light conditions (14 h light/10 h darkness). The estrous cycle was synchronized using eCG and hCG. During estrous (18 h after hCG injection), the hamsters were naturally mated. After seven days of mating, the hamsters were examined daily for pregnancy diagnosis and aging with an ultrasound scanner equipped with an 8.5-MHZ linear probe. On each day of the experiment, at least one of the pregnant hamsters was euthanized and dissected for macroscopic fetal measurements using a digital caliper.ResultsThe gestational sac and crown-rump length were identified and measured by ultrasonographicly on day 7 of pregnancy and head could be visible after day 10 of gestation. Statistical analysis revealed that the ultrasound estimation of gestational age was significantly correlated with the actual age of the fetus (r = 0.98; p < 0.05).ConclusionsReal-time ultrasound can be used for the diagnosis of pregnancy and estimation of fetal age in golden hamster from day 7 of gestation.

Ultrasound characteristics and pregnancy loss in bovine IVF-derived pregnancies

We compared ultrasound characteristics during the first trimester of Girolando female bovine IVF gestations that reached term or not. Pregnant recipients (n = 35) were followed up by ultrasonography every 6 days from D37 up to D90 or until the fetal heartbeat ceased. Embryonic vesicle diameter (EVD), fetometry parameters crown rump length (CRL) and biparietal diameter (BPD), and fetal heart rate (FHR) were assessed in fetuses from recipients that completed gestational development (n = 30) (Control group) or experienced pregnancy loss (n = 5) (Pregnancy Loss group). Results were analyzed by ANOVA repeated measurements. A detection of embryonic vesicle was not possible at D31 in gestations that did not reach term. CRL was decreased in pregnancy loss group and differed at D37 (16.17 ± 0.30 vs 13.14 ± 0.77 mm, for control and pregnancy loss group, respectively) and D55 (46.13 ± 0.51 vs 41.88 ± 2.74 mm, for control and pregnancy loss group, respectively). BPD values were similar between groups. FHR decreased at D49 (181.52 ± 0.82 vs 164.75 ± 9.29 bpm, for control and pregnancy loss group, respectively) and D55 (175.41 ± 2.19 vs 150.80 ± 15.10 bpm, for control and pregnancy loss group, respectively) in early pregnancy losses. These findings indicate that embryonic vesicle, CRL and FHR monitoring of IVF-derived gestations at days 31, 37 and 55 could be useful to predict an increased risk of pregnancy loss.

Early Pregnancy in Jennies in the Caribbean: Corpus Luteum Development and Progesterone Production, Uterine and Embryo Dynamics, Conceptus Growth and Maturation.

Simple SummaryAn understanding of the basic mechanisms of reproduction in donkeys is essential, for several reasons. Some donkey breeds are threatened or endangered, and efforts to save these species depend on improved knowledge of reproductive processes. In some parts of the world, donkeys continue to be valued for purposes of work, recreation, or even meat or milk production, as well as the breeding of mules, and reproduction is essential to maintain suitable populations. In others, donkey populations have become feral and represent a nuisance or even a danger to human populations, and improved contraceptive methods are required. Whether for enhancing or inhibiting reproduction, species-specific information is valuable. While the mare has been extensively studied, few studies have explored early pregnancy in jennies. Therefore, this study characterized early embryo development and differences in progesterone profile and changes in the corpus luteum between pregnant and non-pregnant jennies.We aimed to characterize early embryo development and changes in corpus luteum (CL) development and progesterone profile in pregnant vs. non-pregnant jennies. Eight jennies were enrolled in the study. In the first two cycles, the jennies were monitored by transrectal ultrasonography and had blood harvested for hormone profile assay. In the third cycle, jennies were bred by a jack of proven fertility. Jennies were then monitored and sampled for up to 30 days of pregnancy. Data were evaluated by random-effects multiple linear regression, and correlations were expressed as Pearson’s correlation coefficient. Progesterone concentration rose rapidly from ovulation (D0) until D7, plateaued until D12–14, then precipitously declined between D14 and 15, remaining low until the next ovulation in non-pregnant cycles. In the pregnant jennies, the progesterone concentration rose to maximal concentrations on D7–11, being higher at this stage than in non-pregnant cycles, then declined gradually up to D30. In all cycles, the volume of the CL increased steadily until D6, when it plateaued in pregnant jennies. For non-pregnant jennies, CL volume decreased slowly from D6 to D11 and then had a faster drop. Uterine tone increased following ovulation, becoming turgid around the day of embryo fixation (D15.0 ± 0.9). An embryonic vesicle (EV) was first detected on D9.3 ± 0.5 (2.4 ± 0.5 mm). The EV remained spherical until D18.6 ± 1.4. The embryo proper was first detected ventrally in the vesicle on D20.8 ± 1.1 and the embryonic heartbeat by D22.0 ± 0.9. The allantoic sac was identified at D24.0 ± 0.9, and at D30, the allantoic sac filled the ventral half of the EV. This study provides evidence that higher cumulative concentrations of progesterone are correlated to size of the EV, and there were changes in the luteal dynamics and progesterone profiles in pregnant vs. non-pregnant jennies.

Successful termination of undesirable pregnancy during first trimester in mare

A seven-year-old Marwari mare that was routinely used in semen collection programme was mismated by a fertile stallion. The mare was active and alert and all the clinical and hematological parameters were observed to be normal at the time of examination of pregnancy. Rectal examination of the mare through ultrasonography on day 15 revealed an embryonic vesicle confirming the pregnancy. Initially, the pregnancy was tried to terminate with prostaglandins and prostaglandins in combination with oxytocin, but, the pregnancy persisted and the levels of progesterone (P4) were observed to be elevated. Manual crushing for pregnancy termination was also tried but proved unsuccessful. Uterine lavage by infusing the weak iodine solution in combination with Ringer’s lactate helped in flushing out the conceptus and the levels of P4 were also observed to decrease after the expulsion and the uterus was also found to be devoidof any fluid. The mare showed uneventful recovery and exhibited regular cycles later on and did not show up any abnormalities in conceiving in the consecutive cycles.

A retrospective study of twin pregnancy management in mares

The aim of the study was to compare three methods of reducing twin pregnancy in mares to maintain a single pregnancy. As multiple pregnancies in mare are always undesirable, early ultrasound diagnosis makes possible management of twin pregnancies and extra embryo removal. In years 2010–2018, 16494 mares were sonographically tested for early pregnancy, finding 868 cases of twins (471 bilateral and 397 unilateral). 260 mares with a confirmed bilateral tween pregnancy were subjected to manual crushing of one embryo and administration of flunixin at a dose of 1.1 mg/kg BW. 186 mares were subjected only to the embryo crushing procedure. 25 mares from this group were on a restrictive diet. In the unilateral twin pregnancy mare group, 62 were subjected to manual embryo reduction with simultaneous treatment with flunixin, 60 had only manual embryonic vesicle crush and 210 had a restrictive diet. Determination of success, measured as the development of a single pregnancy, were monitored 2 weeks after the procedure, between the 50th and 60th day of pregnancy and after the 90th day of pregnancy. In general, warm-blooded mares were more prone to a twin pregnancy, and at the same time, all the procedures used to reduce it to a single pregnancy caused a greater risk of losing both embryos than in the case of cold-blooded mares. The beneficial effect of administering flunixin after manual removal of one embryo on the maintenance of the other has been experimentally proven in both unilateral and bilateral twin pregnancy.

25 Estimating Fetal Age Using Transabdominal Ultrasonic Measurements in Sheep Early in Gestation

Abstract An increasingly common method to detect pregnancy in sheep is transabdominal ultrasonography. Research has shown that transabdominal ultrasonography can be accurately measured between days 33–60 of gestation. Additionally, few studies have investigated the use of ultrasonic measurements as predictors of fetal age via transabdominal ultrasound. A pilot study found that pregnancy detection can be performed as early as 31 days with an accuracy of 100% and reported that embryonic vesicle depth (EVD) could be utilized to estimate fetal age. The purpose of the present study was to further evaluate the ability of transabdominal ultrasonography measurements to predict fetal age. Estrus was synchronized in fifteen crossbred ewes, followed by natural mating over a 24h period. From weeks 3 to 9 of gestation, transabdominal ultrasonography (at 3.5 MHz) was performed twice weekly. During on farm ultrasonography, pregnancy diagnosis and fetal count were recorded, and ultrasonic images were recorded for subsequent measurement of EVD and embryonic vesicle length. Eleven out of 15 ewes were confirmed pregnant from our sample. Using the surveyreg procedure in SAS we predict day of gestation through the independent variables: average embryo vesicle depth (aveEVD), aveEVD2, average embryo vesicle length (aveEVL), and the number of fetuses. The model predicts 82% of the variation in gestational age can be explained by the independent variables. Parameter estimates for aveEVD (P &amp;lt; 0.0001) and aveEVL (P &amp;lt; 0.0286) are positive, indicating that every 1 cm increase in these measurements results in 8.9 and 1.3 day increases in gestational age, respectively. Holding other factors constant, we find embryonic vesicle depth increases at a decreasing rate (aveEVD2, P &amp;lt; 0.001). These results indicate gestational age can be predicted through two embryonic vesicle measurements using transabdominal ultrasonography.

Ultrasonographic Detection of Interstitial Nephritis in a Canine Fetus

Background: A detailed ultrasound examination of the fetal urinary tract as part of prenatal care is important to detect possible abnormalities. Early diagnosis can prevent more serious damage such as loss of kidney function. Interstitial nephritis can compromise renal functionality. Also, acute interstitial nephritis is a frequent cause of acute kidney injury and can become chronic if not treated. Besides, a renal biopsy can help in the diagnosis and at the staging of the disease. Therefore, this report aims to describe the ultrasonographic findings of the renal tissues in a canine fetus affected by interstitial nephritis.Case: Gestational ultrasound evaluations were performed in a female Pug, at the 25th, 45th, and 58th gestational days. Ultrasound examination was performed with a 9.0 MHz linear transducer and a high-resolution digital transducer with a frequency standardized at 17.0 MHz. During the first evaluation, embryonic vesicles were detected and were compatible with the gestational period. At 45 days of gestation, some abnormalities were observed in the kidneys of one of the fetuses, such as increased cortical echogenicity, cortical thickening, loss of corticomedullary ratio (1:1), renal pelvis and ureter dilatation by echogenic content. Biparietal and abdominal diameters were also lower than that from the other fetuses, which did not show abnormalities, being compatible with the estimated gestational age. However, the heart rate was similar among all fetuses. On the 58th day, we observed in the underdeveloped fetus that both kidneys still presented the abnormalities seen previously, in addition to the presence of a cystic structure in the right kidney, in the cranial topography of the cortex. The other fetuses kept showing normal ultrasonographic findings, normal heartbeat, all of these compatible with the ultrasound gestational age. The delivery occurred without complications and only the underdeveloped fetus stillborn. Then, it was submitted to a necropsy where chronic interstitial nephritis was found in the histopathological analysis.Discussion: Adequate prenatal care is important to detect abnormalities in fetuses, in which ultrasound examination allows to access fetal development. The gestation in the bitch seemed to be occurring without complications, but the B mode and High Definition ultrasound techniques were able to identify abnormal development of the fetus' kidneys. Both techniques showed to be secure to the fetuses and the mother, as they are non-invasive and free of radiation. These findings, different from those described as normal for a fetus, reinforce the warning signal for a possible establishment of anomalies consistent with nephropathy. Still, histopathology is important to exclude other diseases and help to define prognosis. Interstitial nephritis can significantly affect perinatal survey, considering that the fetus was stillborn, confirming that the description of such anomalies can help to prevent disease. Since nephritis can be associate with the use of some medicines, the orientation of the owner regarding drug use during bitch pregnancy is essential. Therefore, this is the first report describing renal abnormalities in a canine fetus, which highlights the importance of ultrasound examination for disease identification in prenatal care. Lastly, high-resolution ultrasound (HD) could provide detailed information on abnormalities in the fetal kidney.

Celioscopy and partial salpingectomy in a pregnant bitch

ABSTRACT: This case report presents a celioscopy procedure followed by partial salpingectomy (PS) in a pregnant bitch referred to elective video assisted ovariohysterectomy. Through celioscopy it was visualized the embryonic vesicles and PS was performed after conversion to minioceliotomy. The puppies were born by eutocic delivery without clinical alterations. The case suggested that short and regular pneumoperitoneum is not deleterious to dog fetus and video surgery could be used in pregnant bitches. PS allows the fetus development and promotes bitch neutering.

Gestational echobiometry in brachycephalic bitches using high-definition ultrasonography

ABSTRACT: The objective of the present study was to obtain echobiometric values for the most important canine maternal-fetal tissues using high-definition ultrasonography (HDUS) trying to update the formulas for predicting gestational age. Twelve healthy bitches of brachycephalic breeds weighing 7-13kg and aged one to four years were included. The fetuses were analyzed every day using conventional and high-definition ultrasonography from the eighth day after artificial insemination until the day of delivery, using the ACUSON S2000/SIEMENS ultrasound equipment and specific software. Embryonic and fetal structures; diameter of the embryonic vesicle; length of the embryo; biparietal diameter, abdominal length and kidney height; length of the femur, humerus, scapula, radius, and tibia; thickening of the stomach wall; adrenal length; and cardiac size were measured. These variables were correlated with gestational and adjusted tested according to regression models. The results obtained (P&lt;0.001) for: lateral diameter of the gestational sac (R2=81.8%); length of the embryo (R2=85.7%); biparietal (R2=99.1%) and abdominal diameter (R2=97.2%); thickness of the gastric wall (R2=86.9%); length of the femur (R2=96.6%), radius (R2=97.5%), humerus (R2=96.5%), scapula (R2=95.8%) and tibia (R2=97.3%); kidney length (R2=95.8%) and height (R2=96.0%); adrenal length (R2=89.6%); heart length (R2=93.0%) and height (R2=91.5%) of the canine fetuses showed significant correlation with gestational days. This allowed monitoring fetal growth and estimation of age with high accuracy in different gestational periods.

Magnetic resonance imaging for early pregnancy diagnosis in the laboratory rat.

Pregnancy diagnosis and embryo counting are important end points in reproductive, developmental biology and toxicology studies. The purpose of the present study was to assess the feasibility and efficacy of magnetic resonance imaging (MRI) for early pregnancy diagnosis and embryo counting in the laboratory rat. Female Wistar rats were subjected to whole-body MRI scanning using a 1.5T MRI scanner, employing a isotropic T2-weighted 3D short-tau inversion recovery sequence from day 8 to day 12 post coitum (pc) or without prior mating, under general anaesthesia for pregnancy diagnosis and embryo counting. MRI examination was followed by laparotomy and visual inspection of the uterus to verify MRI findings. By day 8 pc, uterine bulges, characteristic of pregnancy, were depicted as oval-shaped structures of high intensity signal. By day 10 pc, embryonic vesicles were detected at the medial side of the uterine bulges. Pregnancy was diagnosed with 0% false-negative diagnosis and 100% accuracy by day 11 pc, while embryos were counted with 100% accuracy by day 12 pc. In conclusion, MRI proved to be a feasible and reliable non-invasive imaging method of early pregnancy diagnosis and embryo counting in the laboratory rat.