• Nathan Riley, MD

Obgyno Wino Podcast Episode 90 - Prenatal Diagnostic Testing for Genetic Disorders

“All at once, without warning of any kind, I found myself wrapped in a flame-colored cloud...I knew that the fire was within myself. Directly afterward there came upon me a sense of exaltation, of immense joyousness accompanied or immediately followed by an intellectual illumination impossible to describe...I saw that the universe is not composed of dead matter, but is, on the contrary, a living Presence; I became conscious in myself of eternal life...I saw that everyone is immortal; that the cosmic order is such that...all things work together for the good of each and all; that the foundation principle of the world, of all the worlds, is what we call love, and that the happiness of each and all is in the long run absolutely certain."

- Richard Maurice Buck

2018 Zinfandel from Bonterra

PB #162, Published May 2016

Five Pearls

1. 1-in-150 live births are affected by a chromosomal abnormality, 1/2 of early pregnancy losses, and 5% of stillbirths

2. Karyotyping is the most best test. It can identify all aneuploidies including 45,X (Turner Syndrome), 47,XXY (Klinefelter Syndrome), and large genomic rearrangements.

3. Microarray is the most reliable method for diagnosing small rearrangements. It's also the preferred method for analysis in pregnancy loss or stillbirth.

4. CVS can be offered from 10-13 wga; amniocentesis can be offered from 15-20 wga.

5. Risk of transmission of Hep B, Hep C, or HIV from amniocentesis or CVS is low as long as viral load is low.

Note: You may want to review PB #226, Screening for Fetal Chromosomal Abnormalities, before diving into diagnostics!

Fetal genetic disorders are complicated...

- we are talking about functional or structural abnormalities in the genome distinct from those caused by environmental factors

- inheritance patterns are very complex

- results of genetic diagnostic testing are never clear enough to make 100% predictions

- not all genetic disorders can be diagnosed through prenatal testing

- chromosomal abnormalities and single-gene disorders can be diagnosed by tissue analysis

- usually abnormal chromosome number (e.g. aneuploidy) or structure

- aneuploidy may also occur as mosaicism (not all cell lines affected)

- chromosomal structural abnormalities include: deletions, duplications, translocations, and other rearrangements

- large structural chromosomal abnormalities may be identified on karyotype analysis

- tiny microdeletions or duplications require microarray, fluorescence in-situ hybridization (FISH), or other specialized methods for detection

- translocations can involve rearrangement, duplication, or deletion of genomic content

- a "balanced" translocation may have normal phenotype but can be associated with recurrent pregnancy loss

- single gene disorders are extremely rare

- examples: sickle cell anemia, cystic fibrosis, hemophilia, Tay-Sachs

- can be identified using fetal cells

...and relatively common

- 1-in-150 live births are affected by a chromosomal abnormality

- likewise, 2/3 cases of blighted ovum, 1/2 of first-trimester pregnancy losses, and 5% of stillbirths

- affects 5-7% of neonatal or early childhood deaths

- more common in the setting of multiple pregnancy losses and structural fetal abnormalities

- isolated structural birth defects are more common within affected families than the general population (often involve multiple genes under influence of environmental factors) --> generally not diagnosable by specific DNA methods but rather by ultrasound

Note: most of the genetic disorders discussed and diagnosed in OBGYN are encoded by nuclear DNA. Mitochondrial DNA can also carry aberrations. Given the important role mitochondria play in aerobic respiration, tissues that rely on aerobic respiration (e.g. CNS, heart, and muscle) can be critically impacted in the event of a rare mitochondrial disorder. Unfortunately, prenatal diagnosis of mitochondrial DNA mutations and associated clinical outcomes is too challenging to discuss in a practice bulletin...

Diagnostic testing requires embryonic cells/tissue

- two methods for obtaining cells/tissue: chorionic villus sampling (CVS) or amniocentesis; more on these later


- the most basic test: can identify all aneuploidies including 45,X (Turner Syndrome), 47,XXY (Klinefelter Syndrome), and large rearrangements

- requires monoculturing of live cells in a lab setting; takes 7-14 days for cells to propagate sufficiently

- >99% accuracy in diagnosing aneuploides or genomic abnormalities greater than 5-10 megabases (1 megabase = 10 nucelotides)

Note: karyotyping can also be done on fetal tissue in the event of pregnancy loss or stillbirth, but it's less accurate given challenges of culturing cells from dead tissue.



- nerds in white coats create fluorescent-labeled probes designed to attach to specific chromosomes or chromosomal regions, then the number of these specific targets in a sample is counted

- direct preparation of cells capture in interphase is much faster as it doesn't require cultured cells (24-48 hrs), but this method only evaluates for major aneuploides

- probes exist for chromosome 13, 18, 21, X, and Y; 22q11.2 (deletion results in what was previously named "DiGeorge Syndrome")

- cultured cells captured in metaphase of mitosis can be used to identify microdeletions and duplications (takes 7-14 days)

- generally pretty accurate but considered more of a screen than a test given higher than desired false-positive and false-negative results (correlate with ultrasound findings)

Chromosomal microarray

- can identify major aneuploidies along with even smaller aberrations that are too small to be detected by conventional karyotyping

- this includes deleted or duplicated sections of DNA ("copy number variants")

- can do with karyotyping can do apart from detecting triploidies or balanced translocations

- like karyotyping, low-level mosaicism may also be missed

- like FISH, doesn't require cultured cells; in fact, it's the preferred method for pregnancy loss or stillbirth (can use tissue from fetus or placenta)

- turnaround is 3-7 days (advantage over karyotyping)

- another advantage over karyotyping is that aberrations small than 5-10 megabases can be detected

- so if US finds structural abnormalities, chromosomal microarray is a better option as a follow-up test than karyotyping for detecting copy number variants (6% of fetuses will be found to have a chromosomal abnormality through microarray who had otherwise normal karyotype)

Note: If a fetal anomaly is found that is strongly suggestive of a typical trisomy like duodenal atresia, which is characteristic of Trisomy 21 (T21), then it's reasonable to start with conventional karyotyping +/- FISH before moving to microarray.

- microarray is such a fine-tooth comb, that in up to 1.7% of fetuses with normal US and normal karyotype, microarray may identify a pathogenic or likely pathogenic copy number variant


- can be done from 10-13 wga: collecting trophoblastic tissue transcervically or transabdominally

- pro over amniocentesis: obtains viable cells, so FISH and microarray can be used (faster turnover time)

- 0.2% risk of pregnancy loss

- also a risk of limb-reduction defects, but only significant if performed at <10 wga

- 1/3 of patients who undergo transcervical approach experience vaginal bleeding

- <0.5% risk of culture failure, amniotic fluid leakage, or infection


- usually done from 15-20 wga (but can be done much later): collecting amniocytes

- risk of alloimmunization if mom is Rh negative (assuming partner Rh positive), though this risk is reduced by avoiding the placenta

- 0.1-0.3% risk of pregnancy loss

- 1-2% chance of minor vaginal spotting or leakage of amniotic fluid

- 0.1% risk of culture failure (need to repeat procedure)

If mom has hepatitis or HIV, there is a risk of vertical transmission with these procedures

- ...but the absolute risk if probably quite small with amniocentesis (limited data)

- risk of HBV transmission is not increased if viral load is low

- 20x increased risk with high viral load (HBV)

- if HBeAg is present, viral replication is active, and transmission risk is higher

- risk of HCV transmission also seems to be low (data also limited)

- as long as your patient is on combination anti-retroviral therapy (CART), risk of transmission is low (esp if viral load is low); reasonable wait until treatment decreases viral load sufficiently before performing procedure

- with CVS, risk is also low (perhaps higher risk with multiple infections)

And let's not forget pre-implantation diagnostic testing...

- uses only a few cells from the early embryo

- can test for specific genetic disorders

- errors can still occur, so confirmation by CVS or amniocentesis is usually recommended if results are concerning

All patients - regardless of age or risk factors - should be offered prenatal assessment for aneuploidy by screening or diagnostic testing

- discuss the differences between screening and testing

- review risk factors for genetic abnormalities based on maternal age, family history, history of genetic abnormalities in prior pregnancies, and ultrasound findings (when appropriate)

- the earlier that this assessment takes place, the better, because management options are more easily digestible for some patients at earlier gestational ages

Risk factors for genetic disorders

Advanced maternal age

- defined as35 years

- higher risk for aneuploidy

- not at higher risk for structural chromosomal anomalies including microdeletions and duplications

Advanced paternal age

- defined as dad aged >40-50 years

- higher risk for single-gene disorders like achondroplasia, Apert syndrome, and Crouzon syndrome

Parental carrier of chromosome rearrangement

- parents who have a translocation or inversion typically have a normal phenotype, but these rearrangements can lead to unbalanced chromosomes and anomalies in offspring

- sometimes this leads to nonviable conception or early miscarriages, so observed risk of live birth with genetic disorder is lower than theoretical risk

- if past pregnancy was affected by an abnormality and a parent is identified as having a chromosomal rearrangement, birth of a child in the future having an abnormality is 5-30%

Parental aneuploidy or aneuploidy mosaicism

- women with T21 have higher risk of offspring with a trisomy

- women with 47,XXX or men with 47,XXY are not at higher risk of having offspring with a trisomy

Prior child with structural birth defect

- conditions such as heart defects or neural tube defects (NTD) tend to recur in families

- risk of recurrence in future pregnancies is 2-3%, dependent on sex of fetus, the specifics of the anomaly, and the number of family members affected

Parenteral carrier of a genetic disorder

- e.g. sickle cell disease, Tay-Sachs, of cystic fibrosis

- higher risk for having an affected child

- if condition is autosomal dominant (e.g. neurofibromatosis), risk is 50%, although some autosomal dominant disorders may arise through spontaneous mutation

- if no family members have been affected in the past, then molecular testing of the affected child is recommended to guide counseling for future pregnancies

Previous fetus or child affected by autosomal trisomy or sex chromosome aneuploidy

- increased risk of 1.5-8x the maternal age risk depending on the specific autosomal trisomy (also dependent on maternal age at time of initial occurrence)

- history of an autosomal trisomy increases risk for any autosomal trisomy

- recurrence risk is higher for 47,XXX and 47,XXY

- not at risk of recurrence for 45,X or 47,XYY

Structural anomalies diagnosed by US

- increased risk of aneuploidy, copy number variants such as microdeletions, and other genetic syndrome

- this risk is dependent on the type and number of anomalies identified

- e.g. isolated echogenic intracardiac focus is a soft marker for T21; this carries a much lower risk for aneuploidy than if you were to find both duodenal atresia + echogenic bowel + echogenic intracardiac focus

When to offer which testing...

- patients at risk for aneuploidy should be offered CVS or amniocentesis with karyotyping

- patients at risk for a genetic disorder should be offered CVS or amniocentesis with DNA testing for the specific disorder

- karyotype or microarray analysis should be offered anytime CVS or amniocentesis is performed but may not be necessary in low-risk patients

- measuring amniotic fluid alpha fetoprotein may not be necessary when amniocentesis is performed for other indications if US of head/neck appears normal

- if major fetal structural abnormalities is seen on US, CVS or amniocentesis with microarray analysis

- if the particular structural abnormalities is strongly suggestive of a particular aneuploidy (e.g. duodenal atresia), karyotype analysis +/- FISH may be offered before microarray analysis

- if risk of a major aneuploidy is suggested due to serum screening or cell-free DNA screening, amniocentesis with karyotype +/- FISH can be offered

Counseling before and after fetal genetic disorder diagnosis

- risks/benefits of testing are an important part of counseling along with the limitations of testing

- if patient/partner wouldn't do anything with the information, perhaps testing is not warranted

- probably best to refer to a genetic counselor before and/or after testing

- in the event that a genetic disorder may be life-limiting or pose risk of in-utero demise, a perinatal palliative care approach would be delightfully helpful (i.e. multi-disciplinary, serial counseling sessions on risk/benefits/alternatives including physicians, social worker, chaplain, and other specialists and support staff)

What about multiple gestation pregnancy?

- counseling is more complex

- estimates for aneuploidy risk with twin pregnancies tend to be overestimates

- if one twin has aneuploidy, options include expectant management, selective termination of affected twin, or termination of entire pregnancy

- risk to one or any of the pregnancies in multi-fetal pregnancy with amniocentesis is unknown, but estimated to be about 2% (risk with CVS ~1%)

- 1% risk of sampling the wrong twin

- if mononchorionic, sufficient to do karyotyping on a single twin, but US isn't superbly accurate in assessing chorionicity (and in rare circumstances even monochorionic twins can have discordant chromosomal analyses)

Some notes on mosacism

- seen in 0.25% of amniocentesis specimens and 1% of CVS specimens

- contamination with maternal chromosomal material can result in false positive mosaicism

- false positive results can be minimized by discarding first 1-2 mL amniotic fluid collect during amniocentesis and by careful dissection of the chorionic villi from maternal decidua

- mosaicism is higher in CVS sample analyzed directly versus cultured cells

- if mosaicism is present on CVS, amniocentesis is recommended to see if mosaicism is present in amniocytes

- if negative on amniocyte analysis, then it could be mosaicism confined to the placenta

- confined placental mosaicism is unlikely to result in birth defects but does carry increased risk for 3rd trimester growth restriction

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