2. Table of Contents
• Prenatal Screening and Diagnosis Of Neural Tube
Defect
• Prenatal Screening and Diagnosis Of Down Syndrome
and Aneuploidies
• Prenatal and preimplantation diagnosis
4. Introduction
• Neural Tube Defect < NTD> : second most prevalent
congenital anomaly, second to cardiac malformation.
• Significant Roles to prevent NTDs:
– Maternal screening programme
– Sonographic imaging with or without the AF testing
– Folic acid as the prevention of NTDs
5. Incidence and Epidemiology
• Highly variable as
– Ethnics and geographic data
– Folic acid supplement programme
– Prenatal screening programme
Screening Maternal AFP with folic
supplement led to decreased
prevalence of NTDs
6. • The Unites States < By 2006>
– 3: 10000
• Englands and Wales < 1970-1997>
– 1: 10000
Higher in Young age, Hispanic and White
The Highest in the United kingdom China
and Egypt
8. Environmental Factors
• Inadequate Folate*** : the nucleic formation was
disturbed then the cell turnover during the neural tube
closure was blocked.
• Drugs** : Disturb the folic acid metabolism : valproate*
and carbamazepine* , coumadin* ,thalidomide* and
efavirenz*
• Insulin dependent DM: induce hypoxia and high
oxidative stress then disturb the signaling pathways led
to embryopathy and apoptosis of neural tube.
9. Environmental Factors
• Hyperthermia: hot tub , sauna
• Amniotic bands : physically disrupt the normal tube
development
• Obesity
• Geographical—United Kingdom, India, China, Egypt,
Mexico, Southern Appalachian United States
10. Genetic Factors
• Polymorphic variants in genes in the folate pathway
• MTHFR polymorphism : Genetic variant 677C to T
which is the thermolabile enzyme which decrease the
enzyme activity therefore the homocysteine levels
would be higher and the DNA synthesis will be
incomplete.
13. • 95 percent of NTDs develop in the absence of a family
history.
• Recurrence risk*****
– 3-5 percents if a couple has a previously had a child
with either anencephaly or spina bifida
– 10 percents if a couple has two affected
children.
– 5 percents if either parents was born with NTDs
14. NTDs Screening
• Candidates : All should be screened since 90-95
percent of cases occur in low risk pregnancy , especially
the one with risk factors< previously affected fetus ,
positive family history , exposure to the associated
medication , GDM and obesity
16. Ultrasound Examination
• Primary screening test for NTD
• Potentially detect NTDs more than MSAFP
• 18-20 wks of GA to perform the screening since the
second trimester ultrasound increase the detection of
spina bifida 92-95 percent and anencephaly to 100***
percent
17. second-trimester fetuses with open spina bifida
• Frontal bone scalloping—the lemon sign****
• Anterior curvature of the cerebellum with effacement
of the cisterna magna—the banana sign****
18. The lemon sign
• Transventricular plane
• Frontal bone
scalloping—the lemon
sign with
ventriculomegaly.
19.
20. The banana sign
• ******
• Transcerebellar plane
• Image of the fetal head
at the level of the
posterior fossa,
• Anterior curvature of
the cerebellum with
effacement of the
cisterna magna—the
banana sign.
25. AFP
– Synthesized by fetal yolk sac and later by GI tract
and liver
– Function: immunoregulation and intravascular
transport protein : analogous of albumin
– normal concentration gradient between fetal
plasma and maternal serum 50,000:1.
– neural-tube and ventral wall defects, permit AFP
to leak into the amnionic fluid, resulting in
increased maternal serum AFP levels
26. AFP
• Maternal serum AFP Peaks at 28-32 wks inconsequence
of the high fetoplacental permeability
• AFAFP < Amniotic fluid AFP > secreted from kidney to
AF and peaks at GA 12-14 wks and declines until
undetectable level at term
• Fetal plasma AFP peaks at 10-13 wks but no clinical
roles
27.
28. MSAFP testing
• MSAFP could be used at GA of 15 -20 wks
• Result was expressed as multiple of median for each
gestational age which can be compared with
different lab and populations
• The value above 2-2.5 of MOMs , designated for
abnormal result.
• If the value elevation persist after a repetition then a
specialised ultrasound examination is needed for
assessment
29. MSAFP
• 2009 meta-analysis
• The positive predictive value is only 2 to 6 percent
• Sensitivity :
– 90 percents in anencephaly
– 80 percent for spina bifida
30. • The positive predictive value of MSAFP level between
2.5-2.9 MOMs for NTDs is 1.45 percents
• MSAFP level greater than 7 MOMs ,the positive
predictive value of for NTDs is 13.5 percents
31.
32. • false-positive rate of up to 5 percent (black hatched
area)
• false-negative rates of up to 20 percent for spina bifida
(tan hatched area) and 10 percent for anencephaly (red
hatched area).
33. Factors increasing MSAFP***
• GA**: increasing by 15 percents per week during
second trimester , if BPD differs more than 1 week
comparing to the stated age then recalculated
MSAFP must be done.
• Maternal weight **: vary in each women in term of
volume distribution
• Diabetes mellitus**: lower the threshold to 1.5
MOMs
34. Factors increasing MSAFP
• Fetal anomalies** : abdominal wall defect, 89 % in
omphalocele and 100% in gastroschisis
• Multiple gestation** : the concentration is
proportional to the number of fetuses : the upper
limit for twins pregnancy is 4-5 MOMs
• Race: Black women
40. AFAFP and AFAChE
• AFAFP : Amniotic fluid AFP
• AFAChE : Amniotic fluid acetylcholine esterase
• A primary biochemical test performed to diagnose
open NTDs when the MSAFP is positive and ultrasound
is equivocal or normal.
41. • An elevation of both AFP and AChE values in amniotic
fluid suggest an open fetal NTDs with 98 percents
accuracy and a false positive rate of 0.4 percents
• Fetal genetic study if amniocentesis is performed .
42. Fetal MRI
• Ultrasounds with limitations including the
reverberation causing suboptimal visualization of the
brain , maternal obesity and oligohydramnios can
hamper the accurate diagnosis
• Further studies are needed to evaluate the diagnostic
capability of MRI ,considered when the ultrasound is
not optimal
43. Prenatal diagnostic study
• Targeted Ultrasound imaging
• If ultrasound is equivocal, AFAFP and AFAChE is
performed as the diagnostic test
• Amniocentesis for genetic study and AFAFP and
AFAChE
44. Additional Diagnostic study
• Ultrasound Survey the other malformation which is
associated with NTD namely oral cleft ,musculoskeletal
renal cardiovascular were the most commonly found as
the associated anomaly.
• 6.5 percents of NTD with anomalies associated with
chromosome and genetic abnormality
• < Trisomy 18 is the most common>***
45. Unexplained MSAFP Elevation
• Normal specialized sonographic evaluation, with or
without amniocentesis
• Increased risk for various adverse pregnancy outcomes. <
Not used as a screening test >
– fetal-growth restriction
– Oligohydramnios
– placental abruption
– preterm membrane rupture
– preterm birth
– fetal death.
46. Pregnancy management
• ACOG recommends the route of delivery for the fetus with
spina bifida should be individualized.
– cesarean section to reduce the chance of mechanical
trauma and spinal infection
– Fetal surgery to improve the motor outcomes but
increasing the maternal and fetal risk.
• Pregnancy termination
– In 83 percents of anenchephaly
– In 63 percents of spina bifida
47. Prevention
• 0.4 mg of folic acid orally every day before
conception and through the first trimester, to reduce
the NTD risk by 80 percents in low risk women****
• 4 mg folic acid taken daily
1 month before ******conception and through
the first trimester in women increased risk for NTDs
• Increased risk for NTDs:
– one or more prior affected children
– either parents has such a defect
48.
49. • Folic acid supplementation may not decrease the risk
for NTDs
• valproic acid exposure
• pregestational diabetes
• first-trimester fever or hot tub exposure
• defects associated with a genetic syndrome
51. Introduction
• Down syndrome is the most common chromosome
abnormality
• Moderate to severe learning disability, heart defect,
intestinal malformation, vision and hearing loss
• High financial and psychological support to the
affected family
• Aneuploid conceptuses
– 50 percent of first-trimester abortions
– 5 to 7 percent of all stillbirths and neonatal
deaths.
52. Prenatal diagnosis and screening test of aneuploidies.
Screening test< noninvasive>
• First trimester
– Ultrasound < nuchal
translucency and CRL>
– Maternal serum marker
< BhcG, PAPP-A>
• Integrated test < Full>
• Sequential test
• Contingent test
• Second trimester
– Triple test
– Quad test
– Maternal serum free cell fetal
DNA
Diagnostic test
• Invasive procedure
– Chorionic villi sampling
– Amniocentesis
– Fetal blood sampling
• Preimplantation diagnostic
testing
53. Candidates for invasive prenatal diagnosis
• Maternal age : risk of having down syndrome and any
aneuploidies increased with age from 35 years to 40
years by four folds comparing to the young age and 10
fold increased risk at the age of 40- 45 .
BUT 70 percent of Down syndrome
pregnancies are in women younger than
35 years.
54. Candidates for prenatal diagnosis
• Dizygotic twins and maternal age older than 31 years at
delivery
• Previous autosomal trisomy , triploidy or other
chromosome abnormality birth
• Patient or partner is a carrier of chromosome
translocation or inversion
• Some cases of repetitive early pregnancy loss
• Major structural defect by sonography
55. First trimester Screening
• Most common used protocol
• GA 11-14wks
– Nuchal translucency alone
– Combined test involoves
• Nuchal translucency and gestational age combines
with serum marker PAPP-A and free or total BhCG
< double test>
56. Nuchal translucency
• This is the maximum thickness of the subcutaneous
translucent area between the skin and soft tissue
overlying the fetal spine at the back of the neck
• It is measured in the sagittal plane, when the crown-rump
length measures between 38 and 84 mm.
59. • The NT measurement is expressed as a multiple of
the gestational age specific median.
• NT ≥ 3.5 mm , offered targeted sonography, in
addition to fetal karyotyping
• One third of fetuses with increased nuchal
translucency thickness will have a chromosome
abnormality
60. • isolated marker, NT detects 70 percent of fetuses with
Down syndrome at a false-positive rate of 5 percent
• maximal sensitivity at 11 weeks
• NT is used as an isolated marker in multifetal gestations
as serum screening is not as accurate or not available.
61. Serum analytes in first trimester
• free β-hCG and PAPP-A <pregnancy-associated plasma
protein A >
• fetal Down syndrome :
– free β-hCG level ≥ 2.0 MoM
– PAPP-A level ≤ 0.5 MoM.
With trisomy 18 and trisomy 13, levels of both
analytes are lower
62. Combined first trimester testing
• NT measurement with serum hCG and PAPP-A.
• Down syndrome detection rates is 79 to 87 percent, a
false-positive rate of 5 percent
• trisomies 18 and 13 detection rates is 90 percent, 2-
percent false-positive rate
63. Full Integrated test
• Ultrasound measurement of nuchal translucency and
serum analytes at the age of 10 -13 then
• At GA of 15-18 wks AFP,uE3, hCG, and inhibin A
• Detection rate 85-95 percents
• Result withheld until quad test completed
• Lowest false positive test in down syndrome
64. Sequential and contingent testing
• The disadvantages of integrated test is patients have to
wait until the second trimester to perform the
screening test thereby the sequential and contingent
testing develops.
65. Sequential testing
• Performing at first trimester
• first trimester screening test and then quad test :
– 1 % offer diagnostic test after screening
– 99% procede to quad test ,result withheld until
quad test completed
• Using first trimester screening and then offering CVS
only to the high risk one.
• If not a very high risk ,go on to the quad test
66. Contingent testing
• First trimester screen and quad test
• Three risk cut-offs
– 1. women at a very high risk< 1%>, > 1 in 50 , after 1
st trimester testing , CVS should be performed.
– 2. women at a very low risk< 84%> ,< 1 in 2000 , no
additional testing after the first trimester screening
– 3. women at intermediate risk < 15%>, > 1 in 2000,
but < 1 in 50, receiving second trimester test.
67. Quadruplet test
• Best available screening test for women who present
for prenatal care in second trimester
• GA 15-18 wks
• Maternal serum for AFP uE3 hCG and inhibin A
68.
69.
70. Abnormal Serum analyte in Down syndromes
Serum analyte Result
1.Total or Free BhCG ≥ 2
2. PAPPA ≤ 0.5
3. AFP ≤ 0.74
4. Unconjugated estriol ≤ 0.8
5. Inhibin alpha ≥ 1.7
71. Cell free fetal DNA in maternal blood
• Next generation genomic sequencing
• 99.5% detection of trisomy 13 18 and 21 and possibly
sex aneuploid
• False positive rate of 0.5% or less
• Perform 12-20 wks of gestation.
72. Free cell fetal DNA in maternal serum
• Candidates for using cell free fetal DNA testing as a
primary screening in singleton
– maternal age of 35 years and older
– Presence of sonographic finding associated with
fetal aneuploidy
– History of previous pregnancy with fetal trisomy
– Screening positive test
– Patients carry a balanced robertsonian
translocation
73. Free cell fetal DNA in maternal serum
• Limitation : uninformative test, expensive
• High sensitivity and specificity : selecting the one who
will be beneficial for the invasive prenatal diagnosis
74.
75. Soft sign
• Soft signs: Normal variants
• Used in 15-22 weeks of gestation
• Risk increase steeply with the number of markers
identified.
78. • 10 percents will have this soft signs in unaffected
pregnancy
• ACOG recommends the risk adjustment based on
second trimester sonographic markers is concerned
because more fetal losses would result than case of of
down syndrome identified.
82. • Nuchal skinfold
– Transcerebellar view of fetal head from the outer
edge of skull to the outer skin
– A measurement of 6 and greater is considered
abnormal.
– More than 10 fold risk of down syndrome
– Amniocentesis should be performed even found as
an isolated finding.
84. Echogenic intracardiac focus
• Echogenic intracardiac focus < EIF> is focal intracardiac
papillary calcification
• Not a cardiac abnormality
• Usually left sided
• Double the risk of down syndrome if found isolated
• Common in trisomy -13
86. Pyelectasis
• Transient or physiological finding in normal fetus
• Measured on transverse image of the pelvis
• 4mm or greater is found about 2 percents of of fetuses
and double the risk of down syndrome
• Likelyhood of having renal abnormality and further
additional testing should perform at GA34wks
88. Echogenic fetal bowel
• Six fold increased risk of down syndrome
• Appears as bright as bones
• Represents :
– Small amounts of swallowed blood
– Fetal CMV infection
– Cystic fibrosis in fetus
90. Femur and Humeral length
• Short femur and humerus
• Short femur: ≤ 90 percents of expected which
correlated with the BPD measurement
• Short humerus: ≤ 89 percents of expected which
correlated with the BPD measurement
93. First trimester sonographic finding in Down
syndrome
• Absent of fetal nasal bone
• Nuchal Translucency
• Wider frontomaxillary facial angle : flat facial profile
• Tricuspid Regurgitation
• Abnormal ductus venosus flow
– also associated with trisomy 18 and 13
Not routinely used in the US
94. Fetal nasal bone
• Two thirds of fetuses with down syndrome, the nasal
bone is not visible in GA of 11-14 wks
• adequate assessment : 45-degree angle of insonation
with the fetal profile; in the midsagittal plane, with the
tip of the nose and the third and fourth ventricles
visible; and that the nasal bone brightness be greater
than or equal to that of the overlying skin
98. Amniocentesis
• Transabdominal withdrawal of amniotic fluid
• Most common prodedure
• Used to diagnose the fetal genetic conditions
• GA 15-20 wks to be performed
• Assess fetal karyotype
• Takes 7 to 10 days to culture the amniocyte and the
assessment of the karyotype
99. Technique
• Sonographic guidance
• Using 20 to 22 gauze spinal needle
• Initially 1 -2 ml of fluid is contaminated
• 20 ml of AF was collected
• After removing of the needle , puncture site is
observed to check bleeding and fetal cardiac motion is
documented
• If maternal Rh-D negative ,RhoGram is given after the
procudure
102. • AF should be clear
• If blood tinged fluid is detected , transplacental passage
is suspected but not associated with pregnancy loss
• In Multifetal pregnancy : indigo carmine dye is often
injected before removing the first needle as the second
sac entrance with clear fluid would verify the needle in
the second sac.
103. complication
• Pregnancy loss : 1 in 300-500
• Double the pregnancy loss if BMI ≥40 kg/m2
• Twins : 1.8 percents
• Amniocentesis leakage < usually within 48 hrs after the
procedure>
• Chorioamnionitis
104. Early amniocentesis
• Performing at the age of 11 -14 wks
• Less fluid is withdrawn : I ml per each GA
• Higher rate of complication : AF leakage, talipes
equinovarus ****and fetal loss
• ACOG not recommend
105. CVS
• Between 10 -13 wks
• Karyotype and specialized genetic test
• Results are earlier ,allowing safer pregnancy
termination
• Karyotype available in 7 -10 days
106. CVS Technique
• Transabdominal
• Transcervical, both are equally safe
• Transcervical villus sampling is performed using a
specifically designed catheter made from flexible
polyethylene that contains a blunt-tipped, malleable
stylet.
107. CVS Technique
• Transabdominal sampling is performed using an 18- or
20-gauge spinal needle.
• With either technique, transabdominal sonography is
used to guide the catheter or needle into the early
placenta —chorion frondosum, followed by aspiration
of villi into a syringe conta
109. • Relative contraindications:
– vaginal bleeding or spotting
– active genital tract infection,
– extreme uterine ante- or retroflexion
. If the patient is Rh D-negative and unsensitized, anti-
D immune globulin is administered following the
procedure
110. • fetal loss rate following CVS was 2 percent compared
with less than 1 percent following amniocentesis.
• An early CVS was its association with limb-reduction****
defects and oromandibular limb
hypogenesis performed at 7 weeks’ gestation
• When performed at ≥ 10 weeks’ gestation, as is
commonly done today, the incidence of limb defects
does not exceed the background rate of 1 per 1000
111. • Vaginal spotting is common following transcervical
sampling, but it is self-limited and not associated with
pregnancy loss.
• The incidence of infection is less than 0.5 percent
112. limitation of CVS
• chromosomal mosaicism is identified in up to 2
percent of specimens.
• In most cases, the mosaicism reflects confined
placental mosaicism rather than a true second cell
line within the fetus.
• Amniocentesis should be offered, and if the result is
normal, the mosaicism is presumed to be confined to
the placenta.
• Confined placental mosaicism has been associated
with fetal-growth impairment and stillbirth.
113. Fetal Blood Sampling
• cordocentesis or percutaneous umbilical blood
sampling (PUBS).
• It was initially described for fetal transfusion of red
blood cells in the setting of anemia from
alloimmunization
• fetal anemia assessment remains the most common
indication.
114. Fetal Blood Sampling
• Fetal blood sampling is also performed for fetal
karyotype determination, particularly in cases of
mosaicism identified following amniocentesis or CVS.
• Fetal blood karyotyping can be accomplished within 24
to 48 hours.
• Although fetal blood can be analyzed for virtually any
test performed on neonatal blood
115. Fetal Blood Sampling Technique
• Under direct sonographic guidance,
• using a 22- or 23-gauge spinal needle into the umbilical
vein, and blood is slowly withdrawn into a heparinized
syringe.
• Adequate visualization of the needle is essential.
• Fetal blood sampling is often performed near the
placental
116. Fetal Blood Sampling Technique
• cord insertion site, where it may be easier to enter the
cord if the placenta is anterior (Fig. 14-9). Alternatively,
a free loop of cord may be punctured.
• a local anesthetic may be administered. Prophylactic
antibiotics are used at some centers, although there
are no trials to support this policy.
117. Fetal Blood Sampling Technique
• Arterial puncture is avoided, because it may result in
vasospasm and fetal bradycardia.
• After the needle is removed, fetal cardiac motion is
documented, and the site is observed for bleeding.
119. • fetal loss rate is approximately 1.4 percent
• Other complications
– cord vessel bleeding in 20 to 30 percent of cases,
– fetal-maternal bleeding in 40 percent of cases in
which the placenta is traversed
– fetal bradycardia in 5 to 10 percent
Most complications are transitory, with complete
recovery, but some result in fetal loss.
120. Preimplantation Genetic Testing
• Genetic testing performed on oocytes or embryos
before implantation ,in vitro fertilization (IVF), may
provide valuable information regarding the
chromosomal complement and single-gene disorders
122. Polar body analysis
• Maternally inherited genetic disorder.
• The first and second polar bodies are extruded from
the developing oocyte.
• Sampling should not affect fetal development
• Disadvantages : paternal genetic contribution is not
evaluated.
123. Blastomere biopsy
• Done at the 6- to 8-cell (cleavage) 3 days old, then one
cell is removed.
• limitation : mosaicism of the blastomeres may not
reflect the chromosomal complement of the developing
embryo
• The technique is associated with a 10-percent reduction
in the pregnancy rate
125. Trophectoderm biopsy
• 5 to 7 cells from a 5- to 6-day blastocyst
• Advantage :no embryal cells are removed as
trophectoderm cells give rise to the trophoblast .
• Disadvantageously :performed later in development
