Equine Pregnancy

Mare Anatomy      

Vulva Vagina Cervix Uterus Oviducts Ovaries

Normal uterus

Period of Embryo • Day 15-55 to 60 - Period of organogenesis. • Maximum mobility from day 11-16. • Fixation between day 14 and 16. • Embryo loses spherical shape, appears triangular at day 17-19. • Amniotic sac begins to form around day 21. • 10% of pregnant mares may show weak estrus and increased follicular activity.

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Fetal heart beat discernible by day 25 on ultrasound exam.

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Day 25 to 30 - If EED occurs, estrus occurs 2-4 weeks later

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Day 28 - Allantois appears and first allantochorionic villi form at yolk sac border

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Early attachment by day 39

• Day 35-40 - PMSG (eCG) – first detectable • Day 42 - change from yolk sac to allantochorionic dependence • Day 50-55 - Accessory corpora lutea formation • Day 55-60 - Villi cover all chorion touching endometrium except endometrial cup areas

Period of fetus • Day 55-60 • Period of progressive increase in fetal crown-rump length • Day 60 - Maximal PMSG concentrations • Day 90 - Estrone sulphate levels begin to increase • Maximum placental attachment occurs by day 150 • Day 80-240 - Maximum growth of fetal gonads and production of estrogens

Rectal palpation

Cervical changes from 16 or 17 d to term are elongation, firmness and tubularity. The uterus also has increased tone.

 

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The chorionic vesicle is distinct and spherical and approximate sizes are: • 28 d (4 wks) Key lime (pullet egg) • 35 d (5 wks) lemon • 42 d (6 wks) orange • 49 d (7 wks) grapefruit • 56 d (8 wks) cantaloupe • By 90 d it is hard to delineate the cranial margin of uterus. • Fetal Ballotment per rectum becomes consistent after 150d. • Aging fetus by size, as in the cow, is imprecise. • Differentials which may confuse the examiner include the bladder and enlargement in the non pregnant tract at the base of the uterine horn.

Rectal examination I. Cervix: • In early pregnancy (16 to 30 days) the cervix can be palpated on the floor of the pelvis as a rigid, firm structure (as during dioestrus).

II. Ovaries: • Both ovaries are usually enlarged from 18 to 40 days due to follicular development and the CL is not palpable. • From 40 to 120 days there is extensive ovarian activity with ovulations, luteinisation and development of secondary corpora lutea. • Follicular activity decreases from 120 days to term and the ovaries become small and inactive for the remainder of gestation. • The position of the ovaries up to 60 days of pregnancy is as for the non-pregnant mare. • From then on they are drawn forward and medially but remain dorsal (above) to the uterus. • From 5 months of pregnancy, the ovaries are not usually palpable.

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Uterus: Both uterine horns and body should be palpated. Pregnancy diagnosis is based on tone and location of uterus and contents. 15 to 22 days The uterus becomes more tubular and turgid from 15 to 22 days post-ovulation and is readily palpable. It can be difficult to feel the conceptual swelling except in maiden mares. 22 to 28 days At about 22 days the conceptual swelling develops at the uterine horn/body junction. This corresponds to fixation, when the conceptus stops migration in the uterus and becomes fixed in position, prior to implantation. The swelling is 3 to 4 cm in diameter (about the size of a golfball) and it bulges ventrally.

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28 to 35 days The uterus is still turgid, and the conceptual bulge is more obvious and the embryonic vesicle is 3 to 6 cm in diameter. The uterine wall over the conceptus begins to feel thin. 35 to 60 days As the conceptus grows the swelling becomes larger and spherical and appears more as a fluid sac. By 42 days it is about 5 x 7 cm (the size of a tennis ball). The uterine tone around the bulge begins to decrease so that the swelling becomes less tense

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60 to 90 days By 60 days the swelling is about 12 cm in diameter and fills the pregnant horn. After 60 days the pregnancy is like an elongated football and starts to involve the uterine body. One must be careful not to confuse it with the bladder or a case of pyometra. The uterus migrates cranially. Between days 60 and 100, the uterus is low within the abdomen and the foetus can not usually be palpated; From 4 to 5 months onwards the foetus can usually be palpated.

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Ultrasound Examination Diagnostic ultrasound plays a pivotal role in the reproductive management of the mare and no deleterious effects of ultrasonography have been reported in man nor in the equine. A thorough understanding of normal ultrasonographic anatomy is vital for veterinarians involved in broodmare work.

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Principles of diagnostic ultrasound • Diagnostic ultrasound utilises sound frequencies between 2 and 10 MHz. • Ultrasound is produced by application of an alternating voltage to piezoelectric crystals which change in size and produce a pressure or ultrasound wave. Returning echoes deform the same crystals which generate a surface voltage. • Most diagnostic ultrasound machines use the principle of brightness modulation (B-mode) where the returning echoes are displayed as dots, the brightness of which is proportional to their amplitude. • Real-time B-mode ultrasound is a dynamic imaging system where information is continually updated and displayed on a monitor. • Ultrasound is attenuated within tissues and attenuation is related to the density of the tissue, the heterogeneity of the tissue and the number and type of echo interfaces. • Bright (specular) echoes are produced when a large proportion of the beam is reflected back to the transducer; these echoes are displayed as white areas on the ultrasound machine screen. • No echoes are produced when the sound is transmitted and not reflected; these areas are displayed as black on the ultrasound machine screen.

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Day 11 pregnancy

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A 14 day conceptus viewed by ultrasound. Note the regularity of the circular shape.

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By 17 days the regularity of shape is starting to deteriorate - note the "ragged" edges now seen on the 17-day pregnancy compared to the 16-day.

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By 23 days the embryo itself can be seen with ultrasound - here it is visible between the red calliper marks on the image. The heartbeat can also start to be detected at this stage. Note here how the regular rounding has completely deteriorated, and the conceptus is appearing oval, and without "smooth" edges.

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At 33 days, the developing allantois (the outgrowth of the embryo's hindgut which forms the bladder, carries blood vessels in the umbilical cord, and later combines with the chorion to form the placenta) is visible, and the yolk sac (visible here as the upper portion of the dark section) is regressing. The apparent "division" in the conceptus at this stage is not to be mistaken for twin embryos - of which clearly only one is visible.

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A different view of a 35 day pregnancy shows that there can be differences Yolk/Allantoic sac. Embryo position.

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By day 39 the yolk sac has almost completely regressed and is visible here only as the dark circular area immediately above the embryo (compare this with the 33 day image).

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In this photograph of a 50day fetus, the legs, head, tail and eye are all clearly defined externally. Internally organs and skeletal structures are in place. Within 10 days or so, the sex of the fetus can be determined using ultrasound, by establishing the direction of migration of the pedicle that will become either the penis or the clitoris. Upward migration towards the anus of course indicates a female.

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Fetal sexing  Numerous reasons for desiring knowledge of the sex of the fetus • Appraisals, • Insurance coverage, • Payment of stud fees, • Sales consignments, • Mating lists, • Sale or purchase, etc.  

Gender determination is based on the location of the genital tubercle.   The genital tubercle is the precursor of the penis in the male or the clitoris in the female.   The tubercle migrates toward the umbilicus in the male and toward the anus in the female. 

Ideal times for performing the procedure are from 59 to 68 days or 5 to 6 months.   Before 58 days the tubercle is not distinct enough and has not migrated sufficiently to make a distinction.   After 70 days the fetus is hard to reach until it is approximately 3.5 to 4 months of age.   As the fetus gets larger, a transabdominal approach may be preferred.  If cattle are available, it is easier to learn 

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External signs of pregnancy    Although abdominal enlargement is characteristic of pregnancy, it is unreliable as a diagnostic sign.   Ballotment or observed movements of the fetus can often be seen late in gestation.   Mammary changes are quite variable.   Pelvic changes (relaxation of the pelvic ligaments) occur late in gestation but are often difficult to detect.   Cessation of estrus behavior is variable and unreliable.   Some mares will continue to show estrus even when pregnant.

Indirect pregnancy tests  

eCG test

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Endometrial cups in a mare.

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The presence of eCG has been used as a test for pregnancy because it is only found in pregnant mares.   The problem is that it remains elevated after the cups are formed even if fetal death occurs. In house tests are available which makes them attractive in some situations (e.g. miniature horses).  For example, with the Synbiotics test, it is reported that 20% of samples are

Plasma progesterone test  

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Progesterone concentrations in plasma can be measured by radio-immunoassay, Alternatively the enzyme-linked immunosorbent assay (ELISA) tests can be used in a veterinary practice laboratory giving the results more rapidly. At 17 to 22 days post-ovulation, pregnant mares should have progesterone levels above 2 ng/ml . However an incorrect diagnosis of pregnancy can occur in cases of: 1. Prolonged luteal phase, 2. Early embryonic death or 3. Mares with a short luteal phase. Not all mares with high progesterone concentrations are pregnant and high progesterone levels only indicate that luteal tissue is present in the ovary. ·

Equine chorionic gonadotrophin (eCG) 

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Produced by fetal trophoblast cells that invade the maternal endometrium beginning around day 35 to form the endometrial cups. Concentrations of eCG in the blood from approximately 40 days after ovulation and these concentrations usually persist at least until 80 - 120 days after ovulation. False positive results are seen when the pregnancy fails after the endometrial cups have formed and continue to produce eCG. False negative results may be seen if sampling is at an incorrect time and in some mares which can be negative for eCG as early as 70 days after ovulation.

Placental oestrogen determination   

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Serum oestrone sulphate levels are detectable From day 65 of pregnancy onwards It increase to peak around day 200, remaining high until after 300 days. The oestrogens come from the foeto-placental unit especially the foetal gonads which are very large during the later stages of pregnancy. Small amounts of oestrogen are produced by the follicles on the mare’s ovaries. In general, this is a reliable assay and can even be performed on faeces. Also a good indicator of foetal viability.

Cuboni test 

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Estrogens are elevated 150 d to term. It is due to production from fetal gonads.  The Cuboni test, based on fluorescence of urine, is 90% accurate after 100 d, 100% accurate after 150 d. 

Estrone sulfate test 

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Estrone sulfate can be tested for in almost any bodily fluid. In serum there is a sharp rise after 60 d, peak levels by 80 d. Before 60 d a false positive can be obtained due to estrus. In milk a similar pattern is observed, only with lower values. It is considered an indicator of fetal viability after 44 d. In feces, it can be found after 4 mos. Also be found in urine. Commercial tests available "Equi Test - ES"

Determination of Oestradiol-17-b Hormone in Faeces 

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The faeces samples of pregnant and non-pregnant mares contained a mean oestradiol-17 b concentration of 9.39 ± 2.63 and 7.70 ± 2.00 ng/g, respectively in the 4th month of gestation. From the 5th month of pregnancy the oestradiol-17 b concentration in pregnant mares was significantly higher than in non-pregnant mares. Oestradiol-17 b levels were estimated to be 71.20 ± 8.00 and 4.60 ± 2.20 ng/g in pregnant and nonpregnant mares in the 5th month of pregnancy, respectively.

Early Pregnancy Factor 

EPF - two components  EPF-A - Uterine tube  EPF -B - Ovary  Production requires signal from fertilized ovum (ovum factor) released under prolactin presence after sperm penetration.  Appears 4-6 hours  Disappears with fetal death  Non-detectable at 20 days in milk and 30 days in serum  Lateral flow dipstick test  It does not work in the cow, so I have doubts in the mare.