prenatal diagnosis and fetal therapy

PRENATAL DIAGNOSIS AND FETAL THERAPY

By-Dr Mohit Moderator Dr Sunita

PRENATAL DIAGNOSIS-

Its the science of identifying structural or functional abnormalities-birth defects-in the fetus.

FETAL THERAPY-

It is used to improve the intrauterine environment for fetal benefit.

it includes-• Blood product transfusion.• Transplacental medication.• Laser or radiofrequency ablation of vascular anastomoses.• Amnioreduction.• Shunt placement.• Extensive fetal surgery.

Why Prenatal screening/diagnosis ?• All the pregnant women are at risk of carrying a fetus with genetic abnormalities.•Incidence of major abnormality apparent at birth is 2 to 3%.

(callens 5th edi)

•Principal goal of prenatal diagnosis is to supply information to at risk families so they can make informed choice during pregnancy.•Potential benefits of prenatal diagnosis are

1) early reassurance to at risk families if results are normal.

2) Risk information to couples who would not have child without defects.

3) Allowing couples to prepare for the birth of an affected child.

4) Risk information to couples for whom termination is an option.

(From callens 5th edi)

• Background risk depends on maternal age and gestational age at time of evaluation. ( Risk increases with age and decreases with gestational age), so prenatal screening should be an integral part of ANC care, not to be limited to high risk population (ACOG 2007).

Genetic counseling • Clinical Genetics is the discipline concerned with the diagnosis and management of the medical, social, and psychological aspect of hereditary diseases.• Genetic counseling is an integral part of genetic testing /prenatal Testing.• The process of genetic counseling involves components like

1)validation of diagnosis

2)obtaining the family history,

3)estimation of risk of recurrence

4)and by all these helping the family to reach decisions and take appropriate action and follow up.• Nondirective counseling is adopted as standard, In this patients are not told what decision to make with regards to testing and management options but, instead, are provided information and support.

First Trimester Screening

Screening methods-1. Age of mother.

2. Nuchal translucency.

3. Nasal Bone.

4. Free Bhcg + PAPPA-P.

5. NT + Free Bhcg + PAPPA-P(combined test).

6. Ductus venosus sonography.

7. Tricuspid Regurgitation Doppler study.Secondary screening tools

Age• Risk of Downs syndrome and any aneuploidy increases with age.• Women with age 35 years and singleton pregnancy and age of 31 years with twins (dizygotic), put women into high risk group of fetal aneuploidy.

Maternal Age Risk of Downs syndrome Total risk for chromosomal abnormalities

20years 1/1667 1/526

25years 1/1250 1/476

30 years 1/1000 1/384

34 years 1/500 1/238

35 years 1/385 1/192

40 years 1/100 1/66

43years 1/50 1/33

45 years 1/30 1/21

49 years 1/11 1/8

Table Showing risk of chromosomal abnormalities in Liveborn infants.(from callens 5th edi.)

Nuchal Translucency

• Nuchal Translucency(NT) refers to the normal subcutaneous fluid filled space between the back of the neck and underlying skin.

• Single most powerful marker available today for differentiating Downs syndrome from euploid pregnancy in 1st trimester.• NT is not specific for aneuploidy its also increased in

1. CHD

2. Genetic syndrome

3. Abnormal or delayed development of lymphatic. • Time – 10 to 14 weeks ( CRL of 36 to 84mm). (from callens 5th edi.)

• Requirements for accurate NT measurement.1. CRL length between 36-84mm.

2. Good midsagittal section showing facial profile.

3.Clear clarification between the fetal skin and amnion achieved by spontaneous or induced fetal movements.

4.Magnification of image so that fetal head, neck, upper thorax, occupy three-fourth of screen.

5.Fetal head should be in neutral position. Extended head will increase and flexed head will decrease the measurement.

6.Reading should be taken in line of the fetal mandible that usually corresponds to area of widest NT.

Nuchal translucency quality control is maintained by national body like in UK its Fetal Medicine foundation who periodically audit interact with sonologist and accredit them for doing NT.

( Callens 5th edi)

NT usefulness

• NT is single most sensitive marker of aneuploidy chances of abnormality in fetus increases in direct proportion to increase in NT value.

Sr No.

NT Values % of abnormal fetuses

1. 2.5 to 3.4 mm 8%

2. 3.5 to 4.4 mm 29%

3. 4.5 to 5.4 mm 48%

4. >6.5 mm 87%

(F Arias 3rd edi)

•The overall Down’s syndrome detection rate is 64% to 70% at 5% false positive rate. (from Williams 23rd).• There are three major trial conducted for detection rate-

Sr no.

Trial Geographical area

Population studied

Detection rate of Downs syndroem

1. SURUSS trial (sr urine and usg study)

UK General population

63%

2. BUN trial(Biochemistry, USG,

NT)

US High risk group

69%

3. FASTER trial (First and Second trimester

eveluation of risk)

Multicentric prospective

trial

General population

64 to 70%.

(ACOG 2007)

NT usefulness

• The measurement of NT is above 95th percentile in 71.8% of fetuses with Downs syndrome, and in 70.5% of fetuses with other chromosomal defects ( T 18, T 13, Turners, triploidy)

71.8% of fetuses with Downs syndrome, 70.5% of fetuses with other chromosomal defects

5th percentile 95th percentile (From F Arias 3rd edi.)

• when used in combination with free Beta-Hcg and PAPP-A----Detection rate of Downs syndrome is 79% to 87%.

• If NT is ≥ 3.5 mm significant risk of aneuploidy ----appropriate to perform invasive testing..

• NT is also useful marker of cardiac anomalies, diaphragmatic hernia, skeletal dysplasia, genetic syndrome.

First trimester screeningAnd detection rate ( from Williams 23rd)

Strategy Analytes Detection rate at 5% false positive rate

USG for NT Nuchal Translucency 64 to 70%

Combined 1st trimester screening

NT, free beta-hcg, PAPP-A 79 to 87%

( Williams 23rd edi)

NT in multiple gestation.1. NT is single most useful tool in screening multiple

gestation.

2. Maternal serum screening results are misleading

1)Serum markers are approx but not exactly, twice those found in singleton pregnancy so relative contribution from each fetus cant be determined.

2) Single maternal serum value is used to provide information about multiple fetus and there are no clear cut guidelines to individualize the risk.

3) In discordant cases, normal fetus will mask the abnormal marker produced by affected twin.

2. NT measurement in first trimester can evaluate each fetus individually and give risk assessment.

Combined 1st trimester screening

• Pregnancies with Downs syndrome are associated with altered levels of serum free beta hcg and PAPP-A.

Beta HCG

1. In Downs syndrome both free alpha and beta subunits are increased, but free beta subunit( Unique) is used for screening.

2. In Downs syndrome placenta is hypersecretory and immature leading to increased secretion of Beta HCG by syncytiotrophoblast.

3. This increase starts at end of the 1st trimester and continues during 2nd trimester. (used in both trimester).

4. Free beta hCG Levels are higher, MOM concentration is ≥2.0 in affected pregnancies. (from williams 23rd)

5. When used alone detection rate is 33% (from F arias)

PAPP-A1. Its concentrations are lower in pregnancies with aneuploidy,

but difference with normal pregnancies becomes smaller with advance in gestational age.

2. Ideal time 10 to 14 weeks.

3. Levels are reduced, MOM concentration is 0.38 in affected cases.

4. When used alone detection rate is 38%. ( F arias 3rd edi)

these 1st trimester serum markers are largely independent of NT, so combining serum and sonographic screening protocol is more effective strategy then alone.

FOR a 5% false positive rate, the Downs syndrome detection rates using combined 1st trimester test is 79 to 87%.

Nasal Bone (NB)• Absent nasal bone on USG done at 11 to 13 weeks is another marker of Downs syndrome.• Absence of Nasal bone is not related to NT and can be combined in one scan.• Detection rate of Downs syndrome using absent nasal bone is 67%.• When combined with NT detection rate is 90%. (F Arias)• Nasal bone detection requires expertise and strictly following criteria for how to take measurement.• No clear recommendation of its use.• Role as second line screening tool in high risk patients and not as a general population screening tool.

Requirement of accurate NB measurement

1) Midsagittal plane.2) Fetal spine should be

posterior.3) slight flexion at neck4) Head and thorax occupy

whole image.5) Face of transducer should

be parallel to the longitudinal axis of the nasal bone and skin over bridge.

6) Two parallel lines representing the skin over the nasal bridge and nasal bone compose of “equal sign”.

7) If bottom part of = sign is missing, nasal bone is considered absent.

1st Line showing Skin

2nd line –Nasal bone

( Callens 5th edi)

First trimester Ductus venosus sonography. • 1st Trimester Ductus venosus blood flow is an adjunctive test

For fetal aneuploidy screening.• Forward triphasic pulsatile flow is normal. •Reversed flow at time of atrial contraction is abnormal.

Ductus venosus sonography usefulness

•Associated with fetal aneuploidy and fetal cardiac malformation.• 59% to 93% of aneuploid fetus has abnormal ductus venosus flow.( Callens 5th edi)•Ductus venous Doppler is difficult as ductus venosus vessel at 10 to 14 week measures 2mm. So difficult to locate and difficult to obtain accurate flow velocity wave form without contamination of waveforms of neighboring vessel like IVC showing reversal of flow during contractions normally.•As secondary screening tool, to be performed by experienced sonologist.• for modifying the final risk for aneuploidy or to help predict prognosis of fetuses with normal chromosome and increased NT.

First trimester Tricuspid Regurgitation(TR)

• Abnormal tricuspid regurgitation in 1st trimester is associated with fetal aneupoidy.• TR is considered to be present if a regurgitant jet of at least 60cm/sec is noted extending to over half of systole• Down syndrome detection rate during 11 to 13 weeks is 68% alone. •Its Easy to obtain TR waveform in comparison to Ductus venosus waveform. • Not recommended for routine screening, has a role as second line screening test (following an initial high risk NT and serum screen) with detection rate of 92%.

(from Callens 5th edi)

2nd Trimester screening

Second trimester screening include following

•Triple Test• Quad Test• Genetic sonogram• Quad Test + Genetic sonogram

Triple Test• Most commonly used serological test in 2nd trimester.• It consist of measuring

1) MSAFP (maternal serum alpha fetoprotein).

2) Beta Hcg.

3) uE3 (unconjugated estriol).• These three variables are independent predictors of genetic risk and in combination with maternal age generate a patient specific risk of having downs syndrome.• Time – 15 to 20 weeks. ( As before 15 weeks there is close overlap of serum markers value in affected and non affected pregnancies.) • Detection rate 60 to 69%. (from Callens 5th edi)

Triple test

• Triple test screens for following fetal disorders.

Disorders MSAFP uE3 Beta hCG Inhibin A

Open NTD increased No change No change No change

Downs syndrome

decreased decreased increased Increased

Trisomy 18 decreased decreased No change No change

Table showing the pattern of results seen in triple and Quad test screening . (from callens 5th).

ALPHA FETO PROTEIN-• Glycoprotein synthesized by fetal yolk sac in early pregnancy

and later by fetal GIT and liver.• Major serum protein analogous to albumin.• Concentration increases in both fetal serum and amniotic fluid

until 13 week, after which levels rapidly decreases, conversely its concentration in maternal serum increases after 12 week.

• Normal ration is 1,00000 : 1 (fetal to maternal serum) When the fetal serum level is 2 million u/l the corresponding amniotic fluid AFP is 20,000 u/L, and maternal serum level is 20u/l

• Fetal body wall defects uncovered by integument such as NTD, ventral wall defect, permits AFP leak in amniotic fluid and leads to its increased concentration in maternal serum, Further in Downs as fetal liver is immature its production is less and rapid removal from fetal circulation so, its concentration in maternal serum.

( Callens 5th edi)

Conditions associated with abnormal level of MSAFP

ELEVATED LEVELS• Underestimation of gestation age. • Multifetal gestation.• IUD• NTD• Gastroschisis and omphalocele• Low maternal weight• Pilonidal sinus• Esophageal or intestinal obstruction.• sacrococcygeal teratoma.• Urinary obstruction• Renal anomalies- Polycystic kidneys, renalagenesis, congenital nephrosis. ( as high as > 10 MOM)•Congenital skin defects•Cloacal extrophy•Chorioangioma of placenta

• Placental abruption• Oligohydroamnios•Preeclampsia•Low birth weight•Maternal Hepatoma or teratoma

Conditions associated with abnormal level of MSAFP

LOW LEVELS• Obesity• Diabetes• Chromosomal Trisomy.• GTD•Fetal death• overestimated gestational age.

MSAFP usefulness

• MSAFP as a component of triple test should be routinely advised in pregnancy.• Results are expressed in MOM and MSAFP level of 2.0 to 2.5 MOM used as upper limit of normal.• 90% detection rate for anencephaly and 80% for spinabifida at MOM 2 to 2.5 as upper limit. (from Williams 23rd).

• Several factors influence the MSAFP and taken into consideration when calculating AFP MoM

Maternal weight

Gestational Age

Diabetes

Multifetal gestation.

Algorithm for evaluating maternal serum alpha feto protein

Follow up When MSAFP is elevated.

• Amniocentesis and determination of AFAFP (Amniotic fluid AFP) and AChE (acetylcholinestarase).• AFAFP and AChE in combination has a 97% detection rate for NTD.• AChE is more specific to neural tissue then AFAFP. • Detection of AChE in amniotic fluid generally indicate an open NTD.

Although increased AChE levels are also seen in

1) Omphalocele.

2) Gastroschisis.

3) cystic hygroma.

4) fetal hydrops.

Unconjugated Estriol

• uE3 is synthesized in placenta, its concentration is decreased in Downs syndrome independent of AFP and hCG levels.• In downs syndrome primary abnormality involves lack of cholesterol precursor probably due to immature liver function rendering placenta unable to produce uE3.• Women with low uE3 (<0.75MOM) are also at risk of—

1) IUGR.

2) Oligohydroamnios

3) Delivery of small for gestational age infants.

4) Those with very low <0.15 MOM at risk of Metabolic and genetic disorder (like Steroid sulphatase def, kallman syndrome,CAH)

Triple test

• For Downs syndrome and Trisomy 18

1. For downs syndrome, the values of serum markers is converted into patient specific risk, 1:250 is taken as cut off

and detection rate is 60% to 69%. (from F arias)

2. For Trisomy 18, the serum concentration of all three analytes is low and 1:100 is taken as cut off and detection rate is 80%.

Multiple marker screening tests are based on a composite likelihood ratio determined by levels of all analytes. The maternal age related risk is then multiplied by this ratio and we obtain patients risk. A positive test indicates increased risk, but its not diagnostic of Trisomy or other aneuploidy. Conversely, a negative screening test indicates that risk is not increased but doesn’t duarantee normal fetus.

Quadruple (Quad) test• The quad test consist of determining the maternal serum concentrates of

1) MSAFP

2) Beta Hcg

3) Free estriol

4) Inhibin A• Time – 15 to 20 weeks• The addition of Inh A to triple test markers will improves the detection rate of downs syndrome in comparison to triple.• It has detection rate up to 81%• Level of Inh A in range of 1.8 MoM .

Combined 1st and 2nd trimester screening

1) Integrated screening –• combines both 1st and 2nd trimester screening test into single risk.• Highest downs syndrome detection rate- 90% to 96%.• Disadvantage being test results are not available until the second trimester test has been complete.

2) Sequential screening-

This test obviate some of the disadvantages seen in integrated test in this test, result of 1st trimester screening are disclosed to women at highest risk, thus allowing them the option of earlier invasive testing and those at lowest risk can still take advantage of higher detection rate achieved with second trimesterscreening.• There are two testing strategies

1) Stepwise sequential screening-

2) Contingent sequential screening-

• Stepwise sequential screening-• In this strategy women determined to be at high risk ( Downs syndrome risk above predetermined cutoff) after 1st trimester screening are offered genetic counseling and option of diagnostic testing, those at low risk offered 2nd trimester screening and both 1st and 2nd trimester results are used for final risk thus increasing detection rate in low risk women and allows early confirmation in high risk women. • Detection rate is 95%.

Contingent sequential screening-• This strategy classifies women as having High Low and Intermediate risk based on 1st trimester screening.•High risk– CVS Low risk—No further screenig

Intermediate risk—2nd trimester screening.• Number of patient entering in 2nd phase are less.• Detection rate is 88 to 94%.

( From ACOG 2007 )

(From ACOG 2007)

60 to

67 to

90 to

INVSIVE PRENATAL DIAGNOSIS TECHNIQUES

• Pre procedure genetic counseling-

Pre procedure counseling is necessary as it will allow patients to understand their situation allow them to give consent(or withhold) for the procedure.

Following points to be explained--

1. The chance that fetus will be affected.

2. Nature and consequences of disorder.

3. Risks and limitation of procedure.

4. Time required for reporting.

5. Possibility of complications of the procedure such as procedure related fetal loss, failed attempt to obtain a testable sample.

INVASIVE PRENATAL DIAGNOSIS TECHNIQUES

Various Techniques are--

1. Amniocentesis.

2. Chorionic villous sampling

3. Fetal blood sampling.

4. Fetal tissue biopsy.

5. Preimplantation genetic diagnosis.

6. Fetal cells in maternal circulation.

Amniocentesis

TYPES of Amniocentesis

1) Mid trimester.

2) Early

Midtrimester amniocentesis

1. Midtrimester amniocentesis is performed at 15 to 20 weeks of gestation.

2. Prerequisite for midtrimester amniocentesis

1. Pre procedure Genetic counseling.

2. Comprehensive USG determining

A) number of fetuses.

B)Gestational age.

C)Placental localization. (for identifying place of puncture).

D) Also rule out congenital anomalies.

3. ABO RH status of the patient.

•STEPS OF AMNIOCENTESIS-

1. Under all aseptic precautions a 20 or 22-G needle is used, under USG guidance needle is inserted in two rapid steps.

2. Once in amniotic cavity fluid is aspirated 1st ml of fluid is discarded because of possible contamination.

3. Total of 20ml of fluid is aspirated and used for fetal karyotyping and for AFP levels.

4. At end of procedure puncture site is observed with help of USG for any bleeding and fetal cardiac activity is documented at end of procedure.

5. Bloody tap is discarded as it overestimate the AFP levels.

Amniocentesis with multiple sacs.

• Requires separate puncture and analysis of amniotic fluid of each sac.• Indigo carmine dye (0.5 to 1.0cc) mixed with 5cc of AF and injected into 1st sac, allows better identification of 2nd sac. Methylene blue dye is not used as associated with hemolysis in fetus.• Dye injection to be given only in cased where 2nd sac is not identify easily.• Care should be taken not to puncture intervening membrane.

Indications of Amniocentesis• Diagnostic

1. Detection of chromosomal abnormalities.

a. Downs syndrome

b. Trisomy 18, 13.

c. Inborn errors of metabolism.

d. Sex linked disorder – Turners, klinefelters, fragile x syndrome.

e. NTD – by AFAFP levels

2. Fetal lung maturity by measuring ratio of Lecithin and sphingomycelin in later pregnancy.

3. Spectrometric analysis of AF determination of degree of fetal hemolysis in Rh neg mother.

• Therapeutic

1. Decompression of polyhydroamnios.

(From F Aris )

1. Complications

1. Transient vaginal discharge or amniotic fluid leakage in 1 to 2%

2. Chorioamnionitis in <0.1%.

3. Procedure related loss

According to the FASTER trial the procedure related loss rate was 1/300 to 1/700

i.e. 0.6-0.14% but the results were questioned on ground of methodology but further studies showed loss rate of 1/200 to 1/300 (ACOG 2007).

4. Fetal injury 1 in 1000.

5. Miscarriage rate – 0.5%

( Williams 23rd)

Early Amniocentesis• Procedure is performed between 11 to 14 weeks.• Disadvantages as compare to midtrimester amniocentesis

1) Procedure at this gest age is more challenging due to lake of membrane fusion to uterine wall.

2) Higher rate of complications like

1) Club foot (1.3%).

2) Amniotic fluid leakage.

3) Fetal loss (7.6%) According to CEMAT trail( Canadian Early and mid trimester Amniocentesis trial allocated women to early and mid trimester groups and shown procedure related loss 7.6% in early group and discouraged the early amniocentesis.

4) More cell culture failure requiring second procedure.•Recommendations are against the use of early amniocentesis.(ACOG 2007)

ACOG Recommendations(2007) on INVASIVE TESTING

• LEVEL A

1) Early Amniocentesis should not be performed because of higher risk of pregnancy loss and complications compared with traditional amniocentesis.• LEVEL B

1) Amniocentesis at 15 weeks of gestation or later is a safe procedure, procedure related loss is less than 1 in 300 to 500.

2) In experienced individual and centers, CVS procedure-related loss rate may be the same as those for amniocentesis.

LEVEL C

1) Invasive testing should be available to all women, regardless of maternal age.

2) Patients with increased risk of fetal aneuploidy includes

Women with a previous fetus or child with an autosomal Trisomy or sex chromosome abnormality, one major or at least two minor fetal structural defects identified by USG, either parent with chromosomal translocation, inversion, or parental aneuploidy.

3) Non directive counseling before prenatal diagnostic testing does not require a patient to commit to pregnancy termination if the results is abnormal.

Chorionic Villus Sampling (CVS)

• CVS, a biopsy of the developing placenta is performed at 10 to 13 weeks of gestation for indications similar to Amniocentesis.• Advantages over Amniocentesis are

1. Performed earlier in gestation.

2. Early diagnosis and patient can have option of 1st trimester abortion. (more safer then 2nd trimester.)

• Two commonly used approaches

1) Transcervical

2) Transabdominal.• Sample consist of placental tissue (~20 mg).

20 G needle is used in Transabdominal CVS

16 G polyethylene catheter with a flexible, stainless obturater is used in Transcervical CVS

Transcervical Vs TransabdominalTranscervical

• Placenta in lower 1/3rd • Technically simple to perform.• More comfortable to patient.• Relative contraindications-

– Positive N gonorrhea culture– Active genital herpes.– Active bleeding.– Extreme uterine Ante or

Retroflexion.• Fetal loss rate slightly higher then

amniocentesis.• Rupture of membrane ~0.3%• Light Vaginal bleeding ~8%.

Transabdominal

• Placenta in upper 2/3rd.• Difficult specially if

placenta is posterior.• Discomfort is greater.• Fetal loss rate identical to

Amniocentesis ~ 0.7%

Incidence of Limb reduction defects higher if CVS is performed around 7 weeks, if performed by an experienced operator after 10 weeks incidence of limb reduction same as background risk.(WILLIAMS OBSTRICS)

Cytogenetic analysisAmniocentesis

Amniocyte culture over a period of 10 to 14 days.

Amniocytes are arrested during metaphase stage of cell division

Amniocyte harvestation, fixed on slide and stained with dye.

Chromosomes are then examined under microscope.

Total time

Around 21 days

Final Karyotyping report

Figure showing G Banded male karyotype

•Giemsa staining-- producing specific banding pattern called G banding is most widely used technique.

• Each chromosome stains in a characteristic pattern of light and dark bands.• so involvement of any particular band in any chromosomal abnormality can be detected precisely.

Rapid test for Karyotype Analysis

• FISH (Florescence In Situ Hybridization) Allows visualization of small region on chromosome too small to be identified by karyotyping. Results available with in 24 to 72 hrs. ( as compare to karyotyping around 21 days.) as done on interphase state.Fluorescent DNA probes are used with same nucleotide sequence as the stretch of chromosomal DNA of interest. Probe will hybridize at 2 places reflecting homologous nature normally. If probe hybridizes at > 2 places detects trisomy.

FISH interphase view• Probes to Chromosome 21 are RED and for 13 are GREEN •PRSENCE OF THREE RED SIGNALE S INDICATE TRISOMY 21

Chromosome 21

Quantitative Fluorescent PCR

• Another rapid technique.• Uncultured aminocytes from 1 to 2 ml of amniotic fluid are collected and DNA is extracted.• Chromosome specific short tandem repeats of DNA from selected chromosome are then amplified using markers.• Using an automated scanner, the intensity of fluorescence present for each short tandem repeat is quantified and the number of chromosome alleles derived.• QFPCR identifies aneuploidy and is used in some centers as an alternative to FISH.

Fetal Blood sampling( Cordocentesis)

•It is performed for assessment and treatment of confirmed Red cell or Platelet alloimmunization and in evaluation of nonimmune hydrops.• Detecting sever fetal anemia.•Fetal Blood also can be obtained for genetic analysis when CVS or AMNIOCENTESIS results are confusing.•Blood can also be analyzed for

metabolic disordershematological

disordersAcid base analysis.immunological studies.

Fetal blood sampling

• Under USG guidance 22 G spinal needle is used to puncture umbilical vein.• Arterial puncture should be avoided may result in vasospasm and bradycardia.• Complications include

•Cord vessel bleeding- 50%•Cord hematoma-17%• Fetal- maternal hemorrhage- 66% with ant placenta and 17% with posterior placenta.•Fetal bradycardia-3to 12%.•Fetal death-1.7% (Williams 23rd)

Fetal tissue biopsy• There are many genetic conditions for which there is no specific molecular test is available.•Prenatal diagnosis can only be done by biopsy of fetal part under USG guidance.• eg. 1)Muscle biopsy to diagnose muscular dystrophy or mitochondrial myopathy.

2)Skin biopsy for diagnosis of epidermolysis bullosa.

Pre Implantation Genetic diagnosis

• With use of ART (assisted reproductive technologies), Zygotes affected with a severe genetic disorder can be identified so that they are not used for IVF, as a result only unaffected embryos are selected for implantation.• Can diagnose nearly 200 different single gene disorders.•Two different techniques are used

1)Polar body analysis (already in metaphase suitable for FISH)

2)Blastomere biopsy at 6 to 10 cell stage (3 day old)- one totipotent cell is removed)• Recommendations are against use of this technology solely for advanced maternal age, that it does not improve IVF fertilization success, and it may be detrimental. (Williams 23rd)

Fetal cells in maternal circulation

• Fetal cells are present in all pregnant women, although the concentration is very low only 2 to 6 cells per ml.•Isolation of these cells and use in prenatal diagnosis will obviate need for invasive procedure.• Most important thing is fetal cells sorting techniques which uses fetal cell surface markers for sorting fetal cells, but no universal fetal cell marker is yet identified hampering clinical implication of this technology.•Further its not possible to have fetal cells witout maternal cell contamination and isolation of sufficient quantity of cells for use is also difficult.

(ACOG 2007 Practice bulletin)

Thank you

Introduction to Genetic sonogram •It’s a specific targeted examination for fetal aneuploidy, most specific for Downs syndrome, that searches for the presence of

1)Fetal structural anomalies

2)Aneuploidy markers and

3)Abnormal biometry. (Callen’s 5th)• These aneuploidy markers are called as “soft” markers and these are variations in normal anatomy that, except for their relationship to aneuploidy, are unlikely to be clinically significant. (Callen’s 5th)• soft markers are often transient and nonspecific findings which can also occur frequently in euploid fetuses.

Genetic Sonogram

•Sensitivity of genetic sonogram ranges from 84% among women with advanced maternal age to 90% among women younger than 35 years with abnormal triple test.• Performed between 18 to 20 weeks of gestation.• Major advantage is, it reduces the invasive testing rate and can optimize the selection of candidate for invasive testing in order to minimize the procedure related losses of normal fetuses without significantly decreasing detection rate.

(Callen’s 5th)

Candidate for Genetic sonogram Low risk vs. high risk population

• Inappropriate for LOW RISK population

1)High false positive rate of 12 to 15% in high risk population (which is probably more in low risk).

2)Each marker by itself has only low to moderate sensitivity for Downs, when found in isolation it not necessarily increase the risk but increases cost and anxiety in LOW RISK.

3) In LOW RISK patient the prior risk may be so low that

Presence of one marker will not qualify a patient for Amniocentesis.

4) Further the accuracy of soft markers has only been studied in HIGH RISK population and generalizing the result in low risk is not appropriate.

Prenatal sonographic features of trisomy 21

(Callen’s 5th)


(Callen’s 5th)

Most investigated “Six Soft markers” of downs syndrome with likelihood ratio

( Williams 23rd edi)

Second Trimester Ultrasound Markers Practice Guidelines

Normal Ultrasound (No marker present)

Low RiskIncluding 1)Age < 35 and 2)serum screen negative

High RiskIncluding Age 1) Age≥ 35years

or 2) Serum screen positive

or Both

No testing required Downs syndrome risk adjustment (80% reduction in a priori risk)

From (Callen’s 5th)


ONE ISOLATED SOFT MARKER PRESENT ( Except Nuchal Fold or Absent nasal bone)




or Both

No testing required Offer Genetic Amniocentesis



≥ 2 SOFT MARKERS PRESENT OR Thick Nuchal fold OR Absent nasal bone OR structural anomaly




or Both

Offer Genetic AmniocentesisOffer Genetic Amniocentesis


prenatal diagnosis and fetal therapy

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