pregnancy diagnosis in mare

Surgical and Obstetrics Department By: Mohammad Rahawy

PREGNANCY AND ITS DETECTION IN THE MARE

Surgical and Obstetrics Department By: Mohammad Rahawy

Endometrial cups in Mare

Surgery and Obstetrics Department By: Mohammad Rahawy

In the early part of the second month of pregnancy, the endometrial cups are formed.These are discrete outgrowths of densely packed tissue within the gravid horn, derived as a result of the invasion of fetal trophoblast cells into endometrium, where they subsequently give rise to the endometrial cup cells. Usually, there are about 12 cups present at the junction of the gravid horn and body as a circumferential band. The endometrial cups produce pregnant mare serum gonadotrophin (PMSG), which is now referred to as equine chorionic gonadotrophin or eCG.

Surgery and Obstetrics Department By: Mohammad Rahawy

It is first demonstrable in the blood 38–42 days after ovulation, reaches a maximum at 60–65 days, declines thereafter and disappears by 150 days of gestation. eCG has both ‘ (FSH)-like’ and ‘ (LH)-like’ activity, and it is generally assumed that, in association with pituitary gonadotrophins, it provides the stimulus for the formation of accessory CLs and regulates luteal steroidogenesis. These structures start to form between 40 and 60 days of gestation, either as a result of ovulation, in the same way that the CL of dioestrus is formed (32%), or as a result of luteinisation of anovulatory follicles (68%). Because of the presence of the accessory CLs, the progestogen concentrations in the peripheral circulation increase, to reach and maintain a plateau from about 50 to 140 days and then decline. By 180–200 days the concentrations are below 1 ng/ml, and they remain so until about 300 days of gestation, when they increase rapidly to reach a peak just before foaling and subsequently decline rapidly to very low levels immediately after parturition. The main source of progesterone in early pregnancy is the true corpus luteum and the accessory corpora luteum

Surgery and Obstetrics Department By: Mohammad Rahawy

Methods of pregnancy diagnosis in Mare: Basic examination of the reproductive tract consists of: • Visual examination of the tail, perineum and vulva; • Manual palpation of the cervix, uterus and ovaries per rectum; • Visual inspection of the vagina and cervix per vaginam using a speculum; • Manual palpation of the vagina and cervix per vaginam; • Real time ultrasound imaging per rectum. • Endoscopic examination of the vagina, cervix and uterus in some cases.


Surgery and Obstetrics Department By: Mohammad Rahawy

Manual examination per rectum

Due to the lateral position of the ovaries, one-handed rectal examination makes accurate palpation of the right ovary for the right-handed examiner (and vice versa) difficult if the mare is not well restrained. • Wear a glove and use adequate lubricant. • Mare usually resents passage of hand most and then the elbow. • Completely evacuate rectum of faeces, and feel for uterine horns lying transversely in front of the pubis. Follow these laterally to the ovaries which are cranial to the shaft of the ileum. • Always try to have the hand cranial to the structure which is to be palpated to allow sufficient rectum for manipulation.

Surgery and Obstetrics Department By: Mohammad Rahawy

• Do not stretch rectum laterally if tense; do not resist strong peristaltic contractions – otherwise rectum may tear (especially dorsally, i.e. not adjacent to examiner’s hand). • If the rectum is ballooned with air, feel forward for peristaltic constriction and gently stroke with a finger to stimulate contraction. • Ovaries often lie lateral to broad ligament and are difficult to palpate. They must be manipulated onto the cranio-medial aspect of the ligament for accurate palpation. • Uterus is very difficult to palpate in anoestrus, easier during cycles and easiest during early (up to 60 days) pregnancy, due to increasing thickness and tone of the uterine wall. • Cervix is palpated by sweeping fingertips ventrally from side to side in mid-pelvic area. It is easiest to feel during the luteal phase, but more difficult during oestrus and anoestrus.

Surgery and Obstetrics Department By: Mohammad Rahawy

Manual examination per vaginam: • May feel remnants of hymen – occasionally complete. • Vagina dry in luteal phase and anoestrus, moist in oestrus, sticky mucus in pregnancy. • Palpate cervix for shape, size and patency of canal. • May detect adhesions or fibrosis in the cervix. • Do not force finger along cervical canal if there is a possibility of pregnancy. • Mare’s cervix will allow gentle dilation, without causing damage, at all stages of the reproductive cycle. • Manual examination may not be possible if mare’s vulva is sutured excessively tightly.

Surgery and Obstetrics Department By: Mohammad Rahawy

Methods of Pregnancy Diagnosis: 1- Absence of subsequent oestrus; This method is commonly used by stud personnel and owners as an initial screening method. However: • Some mares show estrous behaviour when pregnant, and these mares may be mated, especially if restrained: this may cause embryonic death, if the cervix is opened during coitus – more likely in old or recently-foaled mares. • It is commonly assumed that the mare will be in oestrus 21 days after mating, and this is not necessarily true. Teasing may therefore be too late in either normal or short cycles.

Surgery and Obstetrics Department By: Mohammad Rahawy

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• If the mare returns home after mating the owners may not be able to recognize oestrus. This is especially true when there is no stallion or other appropriate stimulus. • Some mares which return to oestrus after mating may show no signs, especially those with foals (silent oestrus). • Non-pregnant mares may not return to heat, usually due to prolonged dioestrus (4.11) and occasionally due to anoestrus (at the end of the season or during periods of inclement weather). • Non-pregnant mares may occasionally enter lactational anoestrus, especially if foaling in January–March. • Non-pregnant mares may not demonstrate estrous behaviour if they are protective of their foal

Surgery and Obstetrics Department By: Mohammad Rahawy

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2- Clinical examination; • Ovarian palpation contributes little to pregnancy diagnosis as large follicles may be (and often are) present and the CL is not palpable. • Uterine and cervical changes. • At 18–21 days: good uterine tone and a tightly-closed cervix (as assessed per rectum or vaginam) are indicators of pregnancy. • 21–60 days: good uterine tone, swelling at the base of one or both (twins) uterine horns and tightly-closed cervix; all must be present for positive diagnosis


Surgery and Obstetrics Department By: Mohammad Rahawy
• 60–120 days: swelling becomes less discrete, uterine horns become more difficult to palpate and uterine body becomes more fluid filled and prominent. The extension of the broad ligament between the uterine horn and the ovary (the mesosalpinx) is pulled into a tight band. This is often a difficult time for pregnancy diagnosis. Continuity with the cervix helps identification of the uterus. Fetus can sometimes be balloted. • 120 days to term: cervix becomes softer, fetus becomes more obvious. Dorsal surface of uterine body always in reach. Fetus often felt moving after six months.

Surgery and Obstetrics Department By: Mohammad Rahawy

3- Progesterone concentrationsProgesterone concentrations in plasma (or milk) can be measured by: _ Radio-immunoassay: sample must be sent (delivered) to laboratory and the result may take two or more days to obtain; _ Enzyme-linked immunosorbent assay (ELISA) tests: these can be conducted in a practice laboratory and the results are rapidly obtained (horse plasma can be harvested in 30 minutes without a centrifuge). The cost per sample is lowest when many samples are assayed in a batch (standards do not need repeating). • At 18–20 days post-ovulation pregnant mares should have plasma progesterone concentration above 1ng/ml but

Surgery and Obstetrics Department By: Mohammad Rahawy

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Not all mares with high progesterone are pregnant (cf. prolonged dioestrus, early fetal death and mares with short cycles); _ Mistiming of sampling (relative to previous ovulation) will give erroneous results; _ Occasionally pregnant mares have low progesterone concentrations for short periods of time; _ Thorough clinical examination gives cheaper and more complete and accurate information on the mare’s reproductive status

Surgery and Obstetrics Department By: Mohammad Rahawy

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4- Equine chorionic gonadotrophin (eCG): Equine chorionic gonadotrophin (eCG, PMSG) appears in the blood in detectable concentrations at approximately 40 days after ovulation and usually persists until 80– 120 days after ovulation. The hormone is produced by the endometrial cups. The amount of eCG produced varies greatly from mare to mare, and mares carrying multiple conceptuses do not necessarily produce more than those with singleton pregnancies

Surgery and Obstetrics Department By: Mohammad Rahawy

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• Errors in the test are due to: _ Sampling at the wrong time; _ Some mares producing little eCG after 80 days; _ Mares in which pregnancy fails after the endometrial cups form continuing to produce eCG (false positive); _ Possible loss of potency in samples not tested immediately. • eCG can be detected by radio-immunoassay (commercial laboratories), haemagglutination-inhibition test (commercial laboratories and test kit for practitioners) and latex agglutination test (test kit for practitioners

Surgery and Obstetrics Department By: Mohammad Rahawy

5- Placental oestrogens: Placental oestrogens reach peak concentrations in plasma and urine at 150 days, and concentrations remain high until after 300 days. The amount of oestrogen produced is so great that false positives do not occur due to other conditions. False negative results are also very rare after 150 days. Oestrogens are tested for in the urine – free oestrogens produce a color reaction with sulphuric acid. The Cuboni test is the most accurate but involves an extraction procedure using benzene (carcinogen) and acid. The Lunaas test is simpler, uses acids but is sometimes difficult to interpret. Plasma assays for oestrone sulphate are now commercially available.


Surgery and Obstetrics Department By: Mohammad Rahawy
6- Ultrasound examination
The ultrasound transducer is usually held within the rectum in the sagittal (longitudinal) plane during imaging. • The vestibule and vagina lie within the pelvis in the midline; these structures can be imaged with ultrasound but are indistinct. • The cervix is located cranial to the vagina approximately 20 cm cranial to the anal sphincter and can be identified as a heterogeneous, generally hyperechogenic, region with a rectangular outline.

Surgery and Obstetrics Department By: Mohammad Rahawy

The uterus is roughly T- or Y-shaped; therefore when using a linear ultrasound transducer the outline of the uterine body generally appears rectangular (the transducer is in a sagittal plane) whilst the outline of the uterine horns appears circular (the transducer whilst orientated in the sagittal plane is positioned in a transverse plane with respect to the uterine horn). • The uterus has a central, homogeneous, relatively hypoechoic, region surrounded by a peripheral hyperechoic layer. • The echogenicity of the endometrium and the uterine cross-sectional diameter vary during the estrous cycle; during oestrus the diameter increases and the uterus becomes increasingly hypoechoic, with central radiating hyperechoic lines which are typical of endometrial oedema.

Surgery and Obstetrics Department By: Mohammad Rahawy

The proximal uterine horns are of smaller diameter than the uterine body. • The ovaries can be located by tracing the uterine horns laterally. • Various sections of the ovaries are usually examined by rotation of the transducer; sections are usually taken from a medial position, and sequential sections of the ventral, mid, and dorsal portions of the ovaries are examined. • Ovaries usually contain follicles (which are anechoic), and may contain luteal structures (which are relatively echogenic – varying shades of greywhite); the ovarian stroma may be difficult to appreciate since it may be surrounded by these structures, although it is generally hypoechoic in appearance

Surgery and Obstetrics Department By: Mohammad Rahawy

Diagnosis of early pregnancy:The early conceptus can be imaged when there is sufficient yolk-sac fluid to be imaged. The yolk sac appears as an anechoic structure which, in early pregnancy, is spherical. There is usually a small echogenic region on the dorsal and ventral poles of the conceptus; this is a normal ultrasound artifact • From ten days after ovulation the conceptus can be imaged; it appears as a spherical anechoic structure approximately 2mm in diameter. • The conceptus rapidly increases in diameter to reach approximately 10mm in diameter 14 days after ovulation. The outline remains circular (spherical) presumably because of the thick embryonic capsule. • Until day 16 the conceptus is mobile and may be identified either within the uterine horns or the uterine body. This mobile phase is important for the maternal recognition of pregnancy.

Surgery and Obstetrics Department By: Mohammad Rahawy

During pregnancy diagnosis, careful attention to imaging of the entire uterus is required; the transducer should be moved slowly from the tip of one uterine horn to the other, and then caudally towards the cervix. • Trans-uterine migration usually ceases by day 17, and the conceptus becomes fixed in position at the base of one uterine horn. • From day 17 until day 28 the increase in conceptus diameter is slowed. • After fixation the conceptus rotates so that its thickest portion, the region of the embryonic pole, assumes a ventral position. • The uterine wall adjacent to the dorsal pole of the conceptus becomes thickened. • The conceptus generally retains a spherical outline until approximately 17 days after ovulation after which time it may be deformed by pressure from the transducer; it may then appear triangular or flattened in outline.

Surgery and Obstetrics Department By: Mohammad Rahawy

• The embryo may be imaged from approximately 21 days after ovulation when it appears as an oblong-shaped hyperechoic structure adjacent to the ventral pole of the conceptus. • A heartbeat is commonly detected within the embryonic mass from approximately 22 days after ovulation. It appears as a rapidly-flickering motion in the central portion of the embryonic mass. • Growth of the allantois lifts the embryo from the ventral position and the allantois per se may be identified from day 24, when it appears as an anechoic structure ventral to the embryo. • The size of the allantois increases and that of the yolk sac is gradually reduced until at approximately 30 days after ovulation they are similar in volume. • From day 30 onwards it is possible to image the amnion surrounding the developing embryo.

Surgery and Obstetrics Department By: Mohammad Rahawy

• At 35 days after ovulation the embryo is approximately 15mm in length and the allantois is three times the volume of the yolk sac. • By days 38–40 the fetus is positioned adjacent to the dorsal pole of the conceptus. • At day 40 the yolk sac is almost completely absent, and the umbilicus, which attaches to the dorsal pole, can be imaged. • A reliable relationship exists in early pregnancy between size of the conceptus and gestational age