screening for fetal abnormality in first and second trimester

1. Screening for fetal
abnormality in first and
second trimesters
Moderator- Associate Prof. Dr. Hema Pradhan
Assistant Prof. Dr. Kirtipal subedi
Presenter – Dr. Bijay ranabhat

2. Objectives
• To know when to screen pregnant woman for fetal abnormality.
• To detect fetuses at high risk for chromosal errors,structural defects
and growth abnormalities.
• To know the methods of first trimestmer and second trimester
screening
• To identify group of woman at high risk of having affected child and
justify the offer of diagnostic test.

3. Introduction
• Screening is the process of surveying a population using a specific
markers and defined screening cut off levels, to identify the
individuals in the population at higher risk for a particular disorder.
• The purpose of screening test is to provide information whether the
fetus is at increased risk for abnormalities.
• Screening test provides sensitivity, specificity, positive and negative
predictive values.
• With a positive screening result, a diagnostic test is recommended if
the patient wants to know whether the fetus is affected.

4. Goals of screening test
• Provide reassurance and remove anxiety, especially among high risk
group.
• Allow couples at high risk to know the presence or absence of the
disorder can be confirmed by testing.
• Provide earlier diagnosis with the option of earlier termination of
pregnancy.
• Allow the couples the option of appropriate management.
• To enable the prenatal treatment of fetus.

5. Congenital abnormalities
• Congenital abnormalities are found in 3-8% of all fetuses and newborns.
• Numerical abnormalities occur due to the addition or deletion of an entire
single chromosome.( trisomy/ monosomy). Most common are
- trisomy 21 ( 15 per 10000 births)
- trisomy 18 ( 3 per 10000 births)
- trisomy 13 ( 2 per 10000 births
- monosomy x ( 2 per 10000 births )
- XXY( 10 per 10000 births)
• Structural abnormalities occurs due to the deletions (when part of a
chromosome is deleted or missing), duplications (when extra genetic
material is added) and translocations (when part of one chromosome is
transferred to another).

6. • Genetic disorders
- multiple malformation syndrome ( holt oram, craniosynostosis)
- hematological disorder ( thalassemia's, hemoglobinopathies,
hemophilia)
- metabolic disorders ( Tay sach , Wilson, Neiman picks disease)
- neuromuscular disorders ( Huntington chorea, myotonic dystrophy,
fragile x )
- renal disorders ( PKD, MCDK)
- connective tissue disorder( osteogenesis imperfecta, Ehlers Danlos)
- skin disorders (epidermolysis bullosa, ectodermal dysplasia)
• Non genetic fetal disorders : fetal infections, immune hydrops, DM,
fetal effects of maternal drugs ).

7. First trimester screening
• Carried out between 11 and 13+6 weeks of gestation, equivalent to a
crown rump length (CRL) of 45–84mm.
• Done by using
- Maternal age
- Nuchal translucency
- Maternal serum beta-hCG level
- Maternal serum PAPP-A level
- Additional ultrasound markers

8. Maternal age
• Advanced maternal age increases the likelihood of having a pregnancy
affected by a chromosomal abnormality.
• Risk of trisomies 13, 18 and 21 all increase with advancing maternal
age.
• The risk of Turner syndrome is not altered with maternal age.
• The detection rate (DR) of trisomy 21 in a woman over 35, using
maternal age only, is 30% for a false positive rate (FPR) of
approximately 5%.
• Combining maternal age and NT, the DR increases to 75–80%.

10. Nuchal translucency (NT)
• Maximum thickness of the subcutaneous translucent area between the
skin and soft tissue overlying fetal spine at the back of neck .
• Increases with gestational age.
• Increased nuchal translucency associated with
- trisomy 21,18,13 , triploidy and turner syndrome.
- spontaneous fetal loss.
- with normal chromosomes : cardiac defects, diaphragmatic hernia,
pulmonary defects, skeletal dysplasia, congenital infection,
metabolic disorders, rare single gene disorders.

11. • Normal pregnancy- chance of normal birth varies with size of NT
measurement.
NT measurement Chance of normal birth
≤ 3.4 mm 95%
3.5 – 4.4 mm 86%
4.5 -5.4 mm 77%
5.5 – 6.4 mm 67%
> 6.5 mm 31%

12. Criteria for Accurate Measurement of Nuchal
Translucency
• The crown-rump length should be 45–85mm.
• Mid-sagittal view of head and upper thorax, occupying the whole screen,
should be obtained. Appropriate magnification (>70% of image).
• The head should be in neutral position in line with the spine.
Hyperextension falsely increases nuchal translucency measurement and
flexion falsely decreases nuchal translucency measurement.
• The fetal neck skin should be differentiated from the amnion, shown by
fetal movements.
• The widest part of nuchal translucency should be measured
• The callipers for measurement should be placed on the inner borders of the
white lines (skin and skull).
• The nuchal translucency measurement should be repeated and the maximum
reading that meets the above criteria should be used.

14. Follow up fetuses with increased NT

15. Maternal serum B-hCG
• Free β chain of Human Chorionic Gonadotropin. Very high in the
early stages of the first trimester, declines in the late first trimester.
• Beta-hCG is increased in fetuses with trisomy 21 from the end of the
first trimester and throughout the second trimester, making it a useful
serum marker in both the first and second trimesters.
• Decreased in trisomies 18 and 13.
• Ethnicity, weight, smoking status and method of conception can alter
the biochemical markers.

16. Maternal serum PAPP-A
• Homotetrameric glycoprotein synthetized in chorionic villi.
• Specific and potent inhibitor of granulocyte elastase.
• Reduced in the majority of fetuses with aneuploidy.
• PAPP-A at 9 weeks, then performing the NT scan and beta hCG levels
at 12 weeks’ gestation could increase the sensitivity of screening for
Down syndrome to 95%.

17. Serum Beta-hCG and PAPP-A Levels at 11–13
Weeks in Pregnancies Complicated with Trisomies
Beta hCG Median MoM PAPP-A Median MoM
Normal karyotype 1.0 Normal karyotype 1.0
Trisomy 21 2.0 Trisomy 21 0.5
Trisomy 18 0.2 Trisomy 18 0.2
Trisomy 13 0.5 Trisomy 13 0.3

18. Additional markers
• Absence of nasal bone
• Reversed a wave in ductus venosus blood flow
• Tricuspid regurgitation
• When these additional ultrasound markers are combined with maternal
age, NT and serum biochemistry, the DR can be increased from 93%
to 96%, while the FPR is decreased from 5% to 2.5%.

20. Second trimester screening
• Biochemical screening performed at 15-22 weeks .
• Combined with maternal age is not effective as first trimester
combined screening.
• Maternal age and triple test ( B hCG, MSAFP and unconjugated estriol
) have detection rates for trisomy 21 of approximately 60% and 5%
false positive rate.
• Maternal age and quadruple test( B hCG, MSAFP, uE3 and inhibin-A)
have detection rate for trisomy 21 of approximately 80% and 5% false
positive rate.

21. • In twin pregnancies, aneuploidy detection rate are significantly lower.
• Trisomy 18 detection rate is similar to that of down syndrome with a
false positive rate of only 0.5%.
• Quadruple marker screen result was abnormal in 96% of those with
triploidy, in 75% with Turner syndrome, in 44% with trisomy 13 and
in 40% of those with other major chromosomal abnormalities.
• Quadruple marker screening offers no benefit over first trimester
screening, but combining first and second trimester screening yields an
even greater aneuploidy detection rate.

22. MSAFP
• Produced by the liver and gastrointestinal tract of the fetus and
excreted in the urine into the amniotic fluid.
• Reduced by around 25% in cases of trisomy 21.
• Elevated in neural tube and ventral wall defects.
• Using MSAFP level of 2.5 MoM as the upper limit of normal, the
neural tube defect detection rate is at least 90% for anencephaly and
80% for spina bifida with a screen positive rate of 3-5%.

23. Unconjugated estriol(uE3)
• Product of the breakdown in the placenta of dehydroepiandrosterone
sulphate, which is produced by the fetal adrenal glands.
• Decreased by approximately 25% in pregnancies complicated by
trisomy 21.

24. Inhibin A
• Produced by the placenta to inhibit the release of FSH.
• First trimester inhibin A levels are similar in aneuploidy and
unaffected pregnancies.
• Elevated in trisomy 21 to around 1.77 MoM in the second trimester.
• Not used in calculating the risk for trisomy 18.

26. Screening options for fetal aneuploidies
1)Combined screening : NT + first trimester serum markers + MA
Sensitivity- 85% FPR- 5%
Advantages
• Early risk assessment and diagnosis of fetal abnormalities allow
maximum time for decision making.
• Allow safer methods of pregnancy termination.
2) Fully integrated testing: NT + first trimester serum markers + second
trimester markers. Sensitivity- 90% FPR- 2%.
advantage- lower false positive rate
Disadvantage- final test results are not available until the second
trimester.

27. 3) Serum integrated screening : PAPP-A + second trimester serum
markers. 85 % detection rate with FPR 4 %.
4) Sequential screening: a variation of integrated test in which small
proportion of highest risk women (>1:50) are identified as screen
positive in first trimester. The remaining women are provided a risk
after all tests have been completed in the second trimester.

29. Sonographic screening
• Soft markers – sonographic detection of aneuploidy, particularly
Down syndrome may be improved by minor markers collectively
known as soft sigs.
First trimester soft signs:
1. NT measurement
2. Nasal bone assessment
3. Ductus venous flow
4. Tricuspid flow

30. 2nd trimester soft signs
• Aberrant right subclavian artery
• Brachycephaly or shortened frontal
lobe
• Clinodactyly
• Echogenic bowel
• Flat facies
• Echogenic intracardiac focus
• Nasal bone absence or hypoplasia
• Nuchal fold thickening
• Renal pelvis dilatation (mild)
• Sandal gap between first and
second toes
• Shortened ear length
• Single transverse palmar crease
• Single umbilical artery
• Short femur
• Short humerus
• Widened iliac angle

31. Cell free DNA screening
• Identify DNA fragments that are derived from apoptotic trophoblasts
which are placental cells undergoing programmed cell death.
• Can be performed any time after 9-10 weeks of gestation.
• Sensitivity to detect down syndrome -99%, trisomy 18- 97% and
trisomy 13- 92 % with respective FPR of 0.1%, 0.2% and 0.2%.
• When fetal fraction is less than 4% , then cfDNA test is considered to
be a failure.

32. • cfDNA is often used as a secondary screening test in the following
high risk groups.
- Maternal age 35 years or older at delivery.
- Fetal ultrasound finding of soft marker associated with an increased
risk of aneuploidies for trisomies. (diagnostic testing is recommended
if structural abnormalities noted)
- Prior pregnancy with autosomal trisomy.
- A positive first- or second-trimester analyte-based screening test.
- Known carriage (patient or partner) of a balanced robertsonian
translocation involving chromosome 21 or 13.

33. Ultrasonographic diagnosis of fetal anomalies
first trimester
1) Neural tube defects- anencephaly and encephalocele can be
diagnosed on 11- 13+6 wog.
Holoprosencephaly – failure to identify both choroid plexuses (the
butterfly sign) is a first trimester warning sign of holoprosencephaly.
2) Cystic hygroma- singular or multiloculated fluid collection, typically
along the posterior fetal neck and back. Two dilated jugular lymphatic
sacs seen on transverse scan.
- Associated with autosomal trisomy, turner syndrome and cardiac
malformations.

34. 3) Heart defects- prevalence of measure cardiac defects increases with
NT thickness. ( 5.4 per 1000 with NT 2.5-3.4mm to 233 per 1000 with
NT >5.5mm ) .
Screening at 11-13+6 wog : Increased NT, abnormal flow in the ductus
venosus, TR. 58 % detection rate.
Detailed fetal echocardiography at 22-24 wog.
3) Omphalocele – suspected if herniation continues after 12 wog. Most
commonly associated with trisomy 18.

35. 4) Megacystis – longitudinal diameter of bladder 7 mm or more at 10-14
weeks of gestation.
- Associated with 25% aneuploidy rate, mainly trisomies 13 and 18.
- Megacystis between 7-15 mm and the karyotype is normal, 90%
abnormalities will resolve and renal function will be normal.
5) Renal anomalies- bilateral renal agenesis, hydronephrosis and MCDK
have been diagnosed by ultrasound in first trimester.
6)Two vessel cord- can be diagnosed in first trimester , not associated
with increased prevalence of trisomy 21, but increases the risk of
trisomy 18 by sevenfold.

36. Fetal anomaly scan
• Performed at 18-22 weeks of gestation.
• Approximately 50 percent of major fetal abnormalities overall are
detected with standard sonography.
• The sensitivity of detecting anomalies varies on factors such as
gestational age, maternal habitus, fetal position, examination type,
operator skill.
• Maternal obesity has been associated with a 20-percent reduction in
the anomaly detection rate.
• The targeted sonogram is a type of specialized examination. It is
performed when the risk for a fetal anatomical or genetic abnormality
is elevated because of history, screening test result, or abnormal
finding during standard examination.

37. Indications for targeted fetal anatomical ultrasound
examination
• Prior fetus or neonate with a structural or genetic/chromosomal
abnormality.
• Current pregnancy with known or suspected fetal abnormality or confirmed
growth abnormality.
• Increased risk for fetal structural anomaly in current pregnancy
- Maternal diabetes diagnosed before 24 weeks of gestation
- ART to achieve conception
- Maternal prepregnancy BMI >30 kg/m2
- Multifetal gestation
- Abnormal serum AFP or estriol level
- Teratogen exposure
- NT measurement ≥ 3 mm

38. • Increased risk for fetal genetic/chromosomal abnormality in current
pregnancy
- Parental carriage of genetic/chromosomal abnormality
- Maternal age ≥ 35 at delivery
- Abnormal aneuploidy screening test result
- Minor aneuploidy marker
- NT measurement ≥ 3 mm
• Other conditions affecting the fetus
- Congenital infection
- Drug dependence
- Alloimmunization
- Amniotic fluid abnormality

39. Indications for invasive diagnostic testing
• Positive screening test for common trisomies.
• A previous pregnancy complicated by fetal trisomy.
• Family history of genetic disease with a high risk of recurrence in the
fetus.
• Structural anomalies or markers for fetal aneuploidy identified at the
dating scan or fetal anomaly screening.
• Evidence of fetal anemia, including increased MCA PSV and signs of
fetal hydrops.
• Diagnosis of fetal bullous and hyperkeratotic disorders, muscular
dystrophies, metabolic disease and fetal tumors.
• Suspected fetal viral infection and evidence of recent maternal
infection on virology testing.

40. Diagnostic procedures
• Diagnostic procedures used in prenatal diagnosis include:
1. Chorionic villous sampling (CVS)
2. Amniocentesis
3. Fetal blood sampling
4. Fetal tissue biopsy
• Karyotype analysis has diagnosis accuracy of more than 99% for fetal
aneuploidies.
• chromosomal microarray analysis (CMA) is recommended as the first-
line genetic test performed, as it may detect clinically significant
chromosomal abnormalities in approximately 6 percent of fetuses with
normal standard karyotype.

41. Chorionic villous sampling
• Performed between 10 and 13 weeks' gestation.
• The specimen is generally sent for karyotyping or CMA.
• Results are available earlier in pregnancy, permitting more time for
decision-making and safer pregnancy termination, if desired.
Complications
• Procedure related fetal loss (0.2-2%)
• Limb reduction defects if performed before 10 weeks of gestation.
• Risk of confined placental mosaicism.
• Risk of maternal cell contamination.
• Failure to obtain adequate sample.
• Failed cell culture.
• Chorioamnionitis.

42. Technique
• Transabdominal ( preferred)- using 18-20 gauze spinal needle under
ultrasound guidance, chorinic villi aspirated using suction with
syringe.
- 5 to 10 gm sample is sufficient for analysis.
• Transcervical - Transcervical CVS is performed using a specifically
designed catheter made from flexible polyethylene that contains a
blunt-tipped, malleable stylet.

44. Amniocentesis
• Carried out to obtain amniotic fluid for karyotyping from 15 weeks
onwards.
• Early amniocentesis can be performed between 11 and 14 weeks of
gestation but not advisable due to a 1.7% additional risk of fetal loss
when compared to amniocentesis after 15 weeks and increased risk of
talipes.
Complications
- Fetal loss (0. 1 to 0.3 percent). The loss rate may be doubled in women
with BMI >40 kg/m2. In twin pregnancies- 1.8%.
- Chorioamnionitis
- Preterm membrane rupture, oligohydramnios
- Maternal cell contamination.
- Test failure.

45. Technique
• 20-22 G needle should be inserted under continuous ultrasound
guidance.
• The needle is inserted perpendicular to the skin and guided into the
deepest portion of the fluid pocket, avoiding fetal parts and umbilical
cord.
• The initial 1 to 2 mL of fluid aspirate may be contaminated with
maternal cells, it is generally discarded. About 20-30 ml sample is
generally adequate for analysis.

46. Fetal blood sampling
• Also called cordocentesis or percutaneous umbilical blood sampling
(PUBS) .
• Initially performed after 18 weeks but has been successfully conducted
from 12 weeks .
• Indicated in fetal karyotype determination, particularly in cases of
mosaicism identified following amniocentesis or CVS.
• Most common indication- confirmation of fetal anemia and planning
fetal blood transfusion . Confirmation and treatment of fetal
thrombocytopenia.

47. complications
• Procedure related fetal loss/ preterm delivery: 1-2%
• Fetal bradycardia or asystole
• Chorioamnionitis
• Amniotic fluid leakage.

48. technique
• A 20-22 G needle is inserted in preselected site using continuous
ultrasound guidance under aseptic technique.
• 1-2 ml of fetal blood will be adequate for hemoglobin, fetal virology,
or karyotype.
• Arterial puncture is avoided, because it may result in vasospasm and
fetal bradycardia.
Three options for technique
1. Cordocentesis
2. Intrahepatic aspiration
3. Intracardiac aspiration

49. Fetal tissue biopsy
• Most fetal diseases that required tissue biopsy can now be identified
with fetal DNA obtained via CVS or amniocentesis.
• Isolated hepatic liver enzyme activity abnormalities may still require
fetal liver biopsy.
• Some conditions (e.g. Duchene muscular dystrophy ) may have
multiple mutation sites , and it may be useful to aid diagnosis at the
functional, protein level.

50. references
• Cunningham FG, Leveno KJ, Bloom SL, Dashe JS, Hoffman BL,
Casey BM et al. Williams obstetrics. 25th edition. New York: McGraw-
Hill Education; 2018.
• James D, Steer PJ, Weiner CP, Gonik B, Robson SC. High Risk
Pregnancy. 5th edition. University of Cambridge: Cambridge
University Press; 2017.
• Aria’s high risk pregnancy 5th edition