Ultrasonographic Studies on Foetal Development in Sheep by Dr. Ramphal Nain - HTML preview

INTRODUCTION

1-4

II

REVIEW OF LITERATURE

5-15

III

MATERIAL AND METHODS

16-18

VI

RESULTS

19-24

V

DISCUSSION

25-42

VI

SUMMARY AND CONCLUSIONS

43-45

BIBLIOGRAPHY

i-viii

7

LIST OF FIGURES

Fig. No.

Title

1.

Transrectal ultrasonogram of nonpregnant uterus of an ewe

with a 7.5 MHz transducer showing the uterus (Ut) and the

urinary bladder (UB) in a sagittal section with dorsoven-

trally directed beam. The uterus appeared homogenous

coarsely granular close to the anterior border of bladder.

2.

Transrectal ultrasonogram of uterus of an ewe at Day 18 of

pregnancy with a 7.5 MHz transducer. The scan is showing

echoic streak of conceptus (C) in anechoic fluid (F).

3.

Transrectal ultrasonogram of uterus of an ewe at Day 25 of

pregnancy with a 7.5 MHz transducer. The scan is showing

embryonic vesicle (EV) attached to one side of endometrium

(Ed).

4.

Transrectal ultrasonogram of uterus of an ewe at Day 25 of

pregnancy with a 7.5 MHz transducer. The scan is showing

dark image i.e. accumulated uterine fluid (shown by ar-

row).

5.

Transrectal ultrasonogram of uterus of an ewe at Day 25 of

pregnancy with a 7.5 MHz transducer. The scan is showing

anechoic line shown by arrow demarkating the fetus in two

parts.

6.

Transrectal ultrasonogram of uterus of an ewe at Day 32 of

pregnancy with a 7.5 MHz transducer. The scan is showing

echoic embryo (E) surrounded by anechoic embryonic fluid

(F). Embryo with budding (Bu) of limbs (shown by arrows)

is visible.

7.

Transrectal ultrasonogram of uterus of an ewe at Day 39 of

pregnancy with a 5 MHz transducer. The scan is showing

echoic fetus (Ft) surrounded by anechoic embryonic fluid

(F). embryo with budding of head (H), forelimbs (FL) and

hind limbs (HL) is visible (shown by arrows). The

hyperechoic amniotic membrane (A.M.) encircling the em-

bryo is also clear visible.

8

8.

Transrectal ultrasonogram of uterus of an ewe at Day 46 of

pregnancy with a 5 MHz transducer. The scan is showing

cardiac cavity (CC) area, head (H), neck (N), trunk (Tr),

spinal vertebrae (SV), Forelimbs (FL) and mouth parts i.e.

Upper jaw (UJ) and lower jaw (LJ) bones of fully devel-

oped fetus.

9.

Transrectal ultrasonogram of uterus of an ewe at Day 46 of

pregnancy with a 5 MHz transducer. The scan is showing

head (H), bony cage of ribs (R) and spinal vertebrae of fe-

tus (SV). Nodule shaped placetomes (P) were seen.

10.

Transrectal ultrasonogram of uterus of an ewe at Day 46 of

pregnancy with a 5 MHz transducer. The scan is showing

fetus lies ventrally. Head (H) is clear visible. Fetus is sur-

rounded by dotted hyperechoic amniotic membrane (A.M.).

11.

Transrectal ultrasonogram of uterus of an ewe at Day 53 of

pregnancy with a 5 MHz transducer. The scan is showing

placentome (P) i.e. typical halfmoon shaped structure.

12.

Transrectal ultrasonogram of uterus of an ewe at Day 53 of

pregnancy with a 5 MHz transducer. The scan is showing

hyperechoic bilobed genital tubercle (GT) lies in between

the umbilicus (Um) and hindlimbs (HL). Fetus was pre-

dicted as male on the basis of this structure. Bones of hind

limbs are also visible.

13.

Transrectal ultrasonogram of uterus of an ewe at Day 53 of

pregnancy with a 5 MHz transducer. The scan is showing

bifurcation of hooves (shown by arrow).

14.

Transrectal ultrasonogram of uterus of an ewe at Day 60 of

pregnancy with a 5 MHz transducer. The scan is showing

pathological abnormalities i.e. dead fetus (DFt). No normal

skeleton of the fetus, but flickering echoic area in fetal fluid

was seen.

15.

Transrectal ultrasonogram of uterus of an ewe at Day 60 of

pregnancy with a 5 MHz transducer. The scan is showing

coccygeal vertebrae (CV) of tail (T), hindlimbs (HL) bones.

The spinal vertebrae (SV) are also visible with clarity.

9

16.

Transrectal ultrasonogram of uterus of an ewe at Day 60 of preg-

nancy with a 5 MHz transducer. The scan is showing lateral view

of head (H), skull bone (SB), spinal vertebral (SV), optic vesicle

(OV), nasal bone (no), nasal passage (np), buccal cavity (be),

muzzle (mz), lower jaw (LJ) and upper jaw bones (UJ). Fetus is

surrounded- by placentomes (P).

17.

Transrectal ultrasonogram of uterus of an ewe at Day 60 of preg-

nancy with a 5 MHz transducer. The scan is showing large sized

cervical vertebrae of spinal cord (shown by arrow) and fetus is in

sitting position.

18.

Transrectal ultrasonogram of uterus of an ewe at Day 60 of preg-

nancy with a 5 MHz transducer. The scan is showing brainlobes

(shown by arrows).

19.

Transrectal ultrasonogram of uterus of an ewe at Day 66 of preg-

nancy with a 5 MHz transducer. The scan is showing full size

placentome (P) occupying most of the space of uterine lumen.

The hypoechoic central cavity of the placentome (P) is shown by

arrow.

20.

Transrectal ultrasonogram of uterus of an ewe at Day 66 of preg-

nancy with a 5 MHz transducer. The scan is showing genital tu-

bercle (GT) as brightest bilobed hyperechoic structure.

21.

Transrectal ultrasonogram of uterus of an ewe at Day 66 of

pregnancy with a 5 MHz transducer. The scan is showing umbili-

cus (Urn) attached with fetus (Ft).

22.

Transrectal ultrasonogram of uterus of an ewe at Day 73 of preg-

nancy with a 5 MHz transducer. The scan is showing cranial cav-

ity (crc), covered by skull bone (SB), containing brane lobes. The

head (H) of fetus with turning at neck (N), resting on trunk (Tr)

and fetus seems to be in sitting position.

23.

Transrectal ultrasonogram of uterus of an ewe at Day 73 of preg-

nancy with a S MHz transducer. The scan is showing pathologi-

cal abnormalities i.e. dead fetus (DFt). No normal

reflection from the accumulated fluid because of echoic spots

(shown by arrows) in the dark image. The skeleton of fetus is also

smaller than the normal size. The turbidity of the fluid confirms

the dead fetus. No cardinal signs of pregnancy seen.

10

24.

Transrectal ultrasonogram of uterus of an ewe at Day 80 of

pregnancy with a 5 MHz transducer. The scan is showing hori-

zontal sagittal section of mouth. The nasal bone (nb), bones of

lowe jaw (LJ), upper jaw (UJ) and tongue (T) in mouth, eye ball

(eb), fore limbs (FL) are clear visible. The placentome (P) lies

just infront of the mouth.

25.

Transrectal ultrasonogram of uterus of an ewe at Day 90 of

pregnancy with a 5 MHz transducer. The scan is showing the

picture of scrotum (Sc) which is situated in between hind limbs

(HL) and below the tail (T) (shown by arrow).

26.

Conceptus Length

Since the fetal parts were not differentiated because the

obseivation from Day 18 to Day 39 are referred as the value of

conceptus. The conceptus length is increased from Day 18 to

Day 39 or almost linearly. The values at Day 18 were less than

0.8 cm while it reached up to close to 1.6 cm by Day 39.

27.

Conceptus width

The conceptus width also shows linear pattern of growth, the

values reached close to 1.2 cm by Day 39.

28.

Fetal Trunk Diameter

The trunk diameter shows significant increase between Day

46 and Day 53, and between Day 60 and Day 66. The trunk

diameter could not be measured after Day 73 due to large size

of fetus and limitations of transducer.

29.

Fetal Head Diameter

The head diameter shows a typical linear pattern of growth.

The values at Day 39 are close to 1 cm and reached around 5

cm by Day 90.

30.

Crown Rump Length

The crown rump length could be measured only between Day

39 and 53 because of the full fetus was seen during these days.

There was significant increase in crown rump length between

Day 39 and 46 and again significant increase between Day 46

and 53.

11

31.

Fetal Heart Beat

The heart beat ranged between 172 to 155 beats per minute

and there were variations among values. However these were

not significant different (P>0.05). It appeared the values were

higher around Day 25 and Day 32.

32.

Cranial Cavity Diameter

A very good image of cranial cavity containing brain lobes was

obtained between Days 46 to 73. The cranial cavity also shows

almost linear pattern of growth between Day 46 and Day 73.

33.

Internal Uterus Diameter

The internal uterus diameter was measured between Day 18

to Day 39 and later on it became difficult to obtain the full view

of the uterus. The values on Day 18 were close to 1.5 cm that

reached up to over 2.25 cm by Day 39

34.

Placentomes Diameter

The good pictures of placentomes were obtained and diam-

eter were measured on a cross section view. The values were

close to 1 cm on Day 39 and that reached to over 2.5 cm by Day

73 and 2.75 cm by Day 90. There was no significant change

between Day 73 and Day 90

35.

Umbilicus Diameter

The umbilicus diameter was measured between Day 39 and

Day 90. It shows almost linear pattern of increase. However

there was significant change between Day 80 and Day 90.

12

Ch apt er -I

Introduction

Sheep industry plays an important role with its multipurpose utility of meat,

wool, leather products and enriched manure thereby contributing a great deal to-

wards our national economy. The accurate prediction of pregnancy in ewes would

greatly increase efficiency of sheep farming.

The profitability of sheep farming could be improved if a simple and a reli-

able technique is available for the detection of pregnancy. This would enable prompt

remating or culling of non-pregnant ewes, more economical use of supplementary

feeding in late gestation and more accurate planning of production.

For the last many years, various techniques have been used to diagnose the

early pregnancy in sheep such as recto-abdominal rods, vasectomized rams, abdomi-

nal ballotment, serum steroid assays and radiography. Most of these techniques have

been unsatisfactory due to factors such as expenses, low accuracy rates, impractica-

bility, ewe and human safety considerations and long delay in availability of results.

Introduction

The use of ultrasound system has improved both the reliability and practi-

cality of pregnancy diagnosis in the field. A mode (Amplitude mode) ultrasound ap-

plied to the flank region has proven to be reasonably reliable from 50 to 120 days of

gestation (Watt et al. , 1984).

Doppler systems have been used rectally from 25 days but require animal

restraint and operator training (Deas, 1977). Recent improvements in ultrasound tech-

nology now enable instantaneous and continuous images of internal anatomy (real

time) (Kossoff, 1979; Simpson et al. , 1982).

Recently portable real-time B-mode (Brightness mode) ultrasound systems

have been used for pregnancy testing in ewes.

In the non-pregnant ewe or in the pregnant ewe during the first 25 days of

pregnancy the uterus is situated within the pelvic cavity. The urinary bladder is taken

as a landmark to take the image of uterus with ultrasound.

Pregnancy is diagnosed by imaging fluid in the uterine lumen, evidence of

conceptus with heart beat and by the presence of placentomes or by the identifica-

tion of fetuses.

Lindahl (1971) was the first who reported the use of B-mode real-time ul-

trasonography with a 5 MHz rectal probe. In view of this reported improvement in

accuracy, it was decided to examine further the rectal probe method of early preg-

nancy diagnosis.

Fowler and Wilkins (1980) used ultrasonic scanning in ewes and found ac-

curate rapid, safe and practicable means of diagnosis of pregnancy.

14

Introduction

They reported real-time ultrasound system transabdominal in sheep as reli-

able to determine pregnancy and fetal numbers from day 50 post-breeding. The rectal

use of a 7.5 or 5 MHz probe in the ewe could yield better results, because the early

pregnant uterus is in the pelvic canal.

Fredrikson and Swertsson (1986) reported that ultrasonography technique

enable to detect the dead fetus or fetal abnormalities in addition to pregnancy diag-

nosis.

With the advent of B-mode real time ultrasonography it is now possible to

obtain images of reproductive organs and developing fetus in small ruminants such as

sheep (Buckrell, 1988; Gearhart et al. , 1988; Aiumlamai et al. , 1992 and Garcia et al. , 1993).

It is the noninvasive method and considered superior to non-imaging tech-

niques because it is more accurate and enables to veterinarian to detect viability of

conceptus and its dimensions (Logue et al. , 1987 and Buckrell, 1988).

An additional advantage of the ultrasound system is the prediction of

foetal sex by identifying and locating the genital tubercle (Coubrough and Castell, 1998).

In our country investigation about the use of ultrasonography in general

and diagnosis of early pregnancy in sheep in particular are lacking. Keeping in consid-

eration of this fact the present experiment was conducted in Corriedale sheep with

the following objectives :-

15

Introduction

1.

To obtain images of uterus on day 18th of mating to differentiate preg-

nant and non-pregnant uterus.

2.

To study fetal development through weekly recording of images of de-

veloping fetus.

3.

To assess change in fetal heart beat up to 90 days of gestation.

16

Review of Literature

Ch apt er -I I

Review of Literature

In intensive sheep farming, sonographic examination for pregnancy detec-

tion and determination of fetal numbers is applied routinely (Fowler and Wilkins, 1984;

White et al. , 1984 and Davey, 1986). Generally the findings on non-pregnant uterus as

well as the uterus and conceptus during pregnancy are similar in sheep and goats

(Tainturier et al. , 1983a, b).

Rowson and Moor (1966) conducted the study of development of sheep

conceptus during the first fourteen days and found that by Day 13 to 14 the embry-

onic vesicle lied as a 10 cm long tube in the uterine horn ipsilateral to the corpus

luteum of pregnancy. By Day 16 to 18 it extended into the contralateral horn. Eckstein

and Kelly (1977) reported that during early pregnancy in ewes, the trophoblast rapidly

elongated to occupy both horns and body of uterus by Day 20. King et al (1982) found

that on Day 11 trophoblast vesicle began to elongate, by Day 13th vesicle entered the

contralateral horn, by Day 18 both horns were occupied in ewes.

17

Review of Literature

Buckrell (1988) reported that real-time p-mode ultrasonography was found

to be a reliable method for diagnosis of early pregnancy in sheep.

Buckrell et al. (1986) and Gearhart et al. (1988) reported that transrectal

ultrasonography occasionally allowed the visualization of anechoic sections through

embryonic vesicle between Days 14 and 19 of pregnancy in ewes. The expanding vesicle

contained inadequate fluid in form of pockets. The embryo in form of series of vesicles

could usually found immediately cranial or cranioventrally to the urinary bladder.

Garcia et al. (1993) have also reported hypoechoic structures 4 mm in di-

ameter by Day 17 to 19 postbreeding in ewes whereas embryonic fluid and membranes

were observed using a 7.5 MHz transducer on Day 15 by Schrick and Inskeep (1993) in

same species. Bretzlaff et al. (1993) found that transrectal imaging allowed visualiza-

tion of pregnancy as early as 15 days postbreeding in ewes.

Martinez (1998) reported round or oblong anechoic uterine areas (embry-

onic vesicle) of more than 3 mm in diameter in the uterine lumen by Day 18 of concep-

tion.

Foetal heart beat

Fraser and Robertson (1967, 1968) noted that the foetal heart beats were

faster than that of dam (140-200 beats/minute) by ultrasonic foetal pulse detector.

Richardson (1972a,b) reported that there was highly significant correlation of foetal

heart beat with the number of days of

18

Review of Literature

gestation. He also observed the mean value ranged from 224 beats per

minute at 61 to 70 days of gestation to 182 beats per minute at 91 to 100 days of

gestation.

Fukui et al. (1984) examined 192 mature ewes mainly Suffolk breed once

for a single or multiple pregnancy at 61 to 170 days after mating with probe type fetal

pulse detector (Heart tons model UGR-6C; Aloka Co.). Continuous ultrasound was trans-

mitted of 2.25 MHz frequency and 30 mW/cm2 intensity. Pregnancy was diagnosed

when the fetal heart was detected. Correct diagnosis for non-pregnant, single, twin

pregnancies were 76.9, 74.4 and 88.7 per cent, respectively.

Buckrell (1988) also found that foetal heart could be imaged most by Day

26-28, and by Day 30 should be possible in all ewes. At Day 30 when scanning with 5

MHz head in inguinal region in the standing/ tipped ewe heart of fetus could be im-

aged immediately in many cases, but required a prolonged scan (1-3 minutes in oth-

ers).

Shriek and Inskeep (1993) detected the rhythmic pulsation heart beat within

embryonic vesicles on Day 18 or 19 in ewes with transrectal ultrasonography by using

7.5 MHz transducer by keeping ewes in dorsal recumbency in tilting squeeze chute.

Garcia et al. (1993) confirmed the pregnancy by detection of an embryo proper and by

an embryonic heart beat on Days 21 to 34 and later judged against the number of

lambs born to each ewe. Botero-Herrea et al. (1984) and Bretzlaff et al. (1993)

19

Review of Literature

reported the embryo detection in goats at Day 24 to 26 by using a 5 MHz

transducer.

Placentomes development

Kahn (1994) observed that placentomes are the cardinal signs of pregnancy.

Clotete (1939) found that ewes’ placentomes are small nodules on Day 21. At Day 30

the periphery of the nodules begin to form a thin lip around the flat centre. And by

approximately Day 90 of gestation the placentome reaches it maximum weight and

diameter. The maximum size of placentomes reaches at 74 and 91 day of gestation in

sheep and goat, respectively.

Clotete (1939) also reported the largest cotyledons of ewes had a diameter

of about 3 cm. However Amoroso (1952) reported that the foetal growth was not lim-

ited by the placental development early in pregnancy but gradual increase in weight

of placentomes was correlated with a gradual increase in size.

Alexander (1964) found that number and weight of placentomes were quite

variable between the two uterine horns in ewes. Riera (1984) noted that sheep usu-

ally had 60 and 100 while goats had between 160 and 180 placentomes.

Buckrell et al (1986) used linear array real time ultrasound system rectally

to see if the fluid pockets fetus and placentomes could be imaged at Day 20 to25 and

Day 25 to 30, respectively. Pregnant ewes after Day 30 displayed an irregular

nonechogenic region anteroventral to the

20

Review of Literature

urinary bladder. Included in this region were button shaped echoic areas

assumed to be placentomes and a fetus within the amniotic membrane.

Buckrell et al. (1988) also reported that placentomes could be detected by

transrectally ultrasonography with a 5 MHz linear transducer at Days 28 to 30 of gesta-

tion. Placentomes appeared as small echoic areas on the surface of endometrium.

Haibel (1990) imaged the placentomes in cross view as cup shaped

hyperechoic structures, the concave surface directed towards the uterine lumen.

Bretzlaff et al. (1993) found that placentomes were readily observable by transrectal

ultrasonography through at pregnancy.

Samuel et al. (1995) did ultrasonography of abdomen of 5 year old Alpine

goat using 5 and 3 MHz sector scanning probes. The accumulation of fluid was deter-

mined to be within uterus with placental tissues and placentomes in lumen. They found

abnormalities in placenta because placentomes were small and round to oval not of

typical-C shaped. They confirmed the case of hydrops uteri on the basis of historical

and physical findings.

Doize et al. (1997) concluded that placentome size increased rapidly during

the first 70 to 90 days of gestation in ewes and does. But they found that in ewes there

was a poor correlation of placentome size with gestational age. They used P-mode

ultrasonograph with 5 MHz transducer and did transrectal procedure.

21

Review of Literature

Foetal sex determination

The accurate predictability of the sex of fetus can speed up breeding

programmes and allows an early selection of pregnant ewes for certain purposes ac-

cording to the sex of their fetuses. There are several methods of both embryonic and

fetal sex determination including Karyotyping, H-Y antigens, and DNA probes for em-

bryos or cells obtained by aminocentesis. Fetal sex can also be determined by ultra-

sonic examination of fetal structures.

Curran and Ginther (1991) found that ultrasonography could provide not

only the gender diagnosis but also the expected accuracy level associated with the

diagnosis. However, Noden and Lahunta (1985) reported that ultrasonographic fetal

sex determination by identification of the genital tubercle could be done in ewes with

a single transrectal examination. The genital tubercle was the embryonic structure

that differentiated into the penis in males and clitoris in females. During differentia-

tion the tubercle moved from the initial position between the rear legs towards the

umbilical cord in males and towa