Prenatal Screening and Diagnosis for Chromosomal Abnormalities and Neural Tube Defects

The American Congress of Obstetricians and Gynecologists (ACOG) recommends that diagnostic testing for chromosome abnormalities and open neural tube defects (ONTD) be offered to all pregnant women. In women of low to average risk, traditional serum testing is often used as a first tier screen, with cell-free DNA (cfDNA) screening as a second-tier test, although cfDNA is also acceptable as an initial screening test.

Quick Answers for Clinicians

Diagnosis

Indications for Testing

  • Diagnostic testing for chromosome abnormalities and ONTD should be offered to all pregnant women; those at high risk for either a chromosome abnormality or ONTD should have more extensive counseling
    • High risk includes
      • Previous pregnancy with chromosome disorder (eg, Down syndrome [DS]/trisomy 21 [T21], trisomy 13 [T13], trisomy 18 [T18], or Turner syndrome [TS]/monosomy X [MX])
      • Parent who is a known carrier of a chromosomal translocation or inversion
      • Maternal age ≥35 years at estimated date of delivery (EDD)
      • Abnormal ultrasound
      • Increased risk of ONTD due to family history, use of specific medications (eg, valproic acid or carbamazepine), or diabetic status
  • Amniocentesis or chorionic villus sampling (CVS) confirms diagnosis suggested by screening tests
    • CVS – 10-13 weeks
      • Ability to examine fetal chromosomes only; uses placental tissue
      • Not appropriate for women at increased risk for ONTD
      • Small chance of identifying a chromosome abnormality that is restricted to the placental tissue and is not present in the fetus (confined placental mosaicism [CPM])
    • Amniocentesis – ≥15 weeks
      • Ability to examine fetal chromosome, amniotic fluid, alpha-fetoprotein concentration, and acetylcholinesterase, if indicated

Screening

Many options for aneuploidy screening are available. Medical practices choose which screening tests to offer based on provider and patient preference, patient population, and service availability. Most professional guidelines endorse offering cfDNA screening to women at increased risk of having a child with chromosomal aneuploidy but suggest that it is more appropriate to offer low- or average-risk women traditional serum screening first and cfDNA as a second-tier screen.

Biochemical Serum Screening

  • Singleton gestations (±nuchal translucency [NT] measurement)
    • Can be done between 10-24 completed weeks gestation
    • Various combinations of pregnancy-associated serum markers available in both the first trimester (ie, placental protein A [PAPP-A] and human chorionic gonadotropin [hCG]) and the second trimester (alpha fetoprotein [AFP], hCG, unconjugated estriol [uE3], and dimeric inhibin A [DIA]) are analyzed with or without NT measurement performed between 10 weeks 3 days and 13 weeks 6 days
    • Screening may be performed in the first and/or second trimester; NT measurement is required for specific screening tests (see below)
    • ACOG issued revised serum screening (±NT measurement) recommendations in 2016
      • All women, regardless of maternal age, should be offered the option of aneuploidy screening or diagnostic testing for fetal genetic disorders
      • Although maternal age may be helpful in adjusting an individual woman's risk of having a fetus with aneuploidy, it should not be used as the sole determinant of whether aneuploidy screening or diagnostic testing is offered
      • Women seen early in pregnancy should ideally be offered aneuploidy screening that combines first- and second-trimester testing (integrated or sequential)
    • Recommend any of the following screening tests based on patient and physician preference (see Test tab and Prenatal Screening table below for more details about these tests)
      • If a patient presents in the first trimester and an NT-certified sonographer is available
        • First-trimester (combined) screen for women who accept a test with a higher screen-positive rate (SPR), but want a T21/T18 test that is complete in the first trimester
        • Sequential screen for women who prefer lower SPR, closer to that of the integrated test but want to know early if they are high risk and who do not mind returning for a second blood draw in the second trimester to complete the test
        • Integrated screen with NT for women who want the best possible detection rate (DR) coupled with the lowest SPR and who do not mind returning for a second blood draw in the second trimester to complete the test or waiting until the second trimester to receive results
      • If a woman presents in the first trimester and an NT-certified sonographer is not available
        • Integrated screen
          • Can be performed without an NT measurement (serum integrated)
          • Results available in the second trimester
          • Best available biochemical-only screening test for women who present in the first trimester and who are not able to have an NT measurement done for any reason
      • If a low-risk patient presents in the second trimester
        • Quad screen
        • Ultrasound (US)
    • Follow-up
      • If screening results are normal – low risk for T21, T18, and ONTD; no further testing recommended  
        • If estimated date of delivery (EDD) changes at scheduled second trimester US (18-22 weeks) by ≥10 days, contact lab to recalculate screening results EVEN IF ORIGINAL SCREENING TEST WAS NORMAL, as it was not based on accurate gestational age
          • Do not redraw sample to repeat test unless recommended by the laboratory
      • If screening results indicate an increased risk for ONTD (high AFP)
        • US to look for twins, fetal/placental abnormalities, and to confirm dates
          • If EDD changes by ≥10 days (based on second trimester US or ≥7 days based on first trimester US) – contact lab to recalculate screening results
          • If EDD does not change based on US, offer genetic counseling, level II ultrasound, and diagnostic testing
      • If screening results indicate an increased risk for T21 or T18
        • US to look for fetal/placental abnormalities and to confirm dates
          • If screen positive for T21 and EDD changes by ≥10 days (based on second trimester US or ≥7 days based on first trimester US) – contact lab to recalculate screening results
            • If recalculated gestational age is <14 weeks (<15 weeks at some labs) when sample was drawn, redraw sample and repeat test; first test was invalid
          • If screen positive for T18 and EDD changes by ≥14 days (based on second trimester US or ≤7 days based on first trimester US) and second trimester US is unremarkable – contact lab to recalculate screening results
            • NOTE: care must be taken when deciding to recalculate a positive T18 screen; fetuses with T18 can have intrauterine growth restriction (IUGR), and may therefore measure smaller than expected in the second trimester if they are affected
            • Consider offering cfDNA screening and/or amniocentesis even if serum screening recalculates to a normal result
            • If EDD does not change based on the US, offer genetic counseling, level II ultrasound, and either screening by cfDNA or diagnostic testing
              • Choosing secondary screening by cfDNA may delay definitive diagnosis and has the potential to miss some affected fetuses
    • If anatomy ultrasound reveals abnormalities or growth restriction, offering prenatal diagnosis (preferred) or cfDNA in the context of normal biochemical screening results is appropriate  
      • If NT ≥3.5 mm
        • High risk for fetal aneuploidy; additional serum screening is not indicated
          • Offer genetic counseling and either screening by cfDNA, or diagnostic testing by chorionic villus sampling (CVS) or amniocentesis, as well as targeted anatomy survey US and fetal echocardiogram in the second trimester
            • Choosing secondary screening by cfDNA may delay definitive diagnosis and has the potential to miss some affected fetuses
      • If NT ≥2.5 multiples of median (MoMs) after serum screening is reported
        • Second-trimester targeted anatomy survey ultrasound and fetal echocardiogram should be offered even if the screening results suggest a low risk for aneuploidy
    • Sonographers must be certified to perform NT measurements by the Fetal Medicine Foundation (FMF) or the Nuchal Translucency Quality Review Board (NTQR)
    • For more test-specific information, refer to ARUP's First- and Second-Trimester Screening Options
  • Multiple gestations (±NT measurement)
    • Screening in twin pregnancies
      • In the case of twins, it is not possible to determine the relative contribution of each fetus to the measured concentrations of the serum markers
      • “Pseudo-risk” for T21 or T18 can be determined by dividing the measured marker by the median value observed in twin gestations and calculating the risk as if the woman were carrying a single fetus
        • Decreases DR as compared to screening women with singleton gestations
        • Accurate NT measurements of both fetuses in the first trimester can increase DR for first trimester screens
      • If twins are monozygotic, they are usually concordant for chromosome abnormalities; if twins are dizygotic, they are usually discordant
    • Screening in higher order multiple gestations
      • Not recommended for T21 and T18 – risk estimates are generally not available
      • Screening for ONTD is available in twin and triplet pregnancies, but DR is decreased

cfDNA Screening

  • Noninvasive prenatal screening tests for T21 and other numerical chromosome abnormalities (chromosomes evaluated depend on the laboratory performing testing)
    • Option of analysis for some relatively common microdeletion syndromes is available through some laboratories
  • Testing is performed on the small fraction of genomic cfDNA that circulates in the maternal bloodstream; this DNA is derived from both mother and fetus (placenta)
  • Testing can be performed as early as 9-10 weeks gestation
  • Testing is available for
    • T21
    • T18
    • T13
    • MX
    • Sex chromosome trisomies (SCTs) XXX, XXY, XYY
    • Microdeletion syndromes
      • 22q11.2 (DiGeorge/velocardiofacial syndrome)
      • 1p36
      • Prader-Willi syndrome
      • Angelman syndrome
      • Cri-du-chat syndrome
    • Some labs also test for Wolf-Hirschhorn, Langer-Giedion, and Jacobsen syndromes; trisomies 16 and 22; or triploidy  
  • Recently, the American Congress of Obstetrics and Gynecology (2016) and the  American College of Medical Genetics and Genomics (2016) stated that cfDNA was the most sensitive screening test for fetal aneuploidy and should be discussed as an option with all pregnant women; however, the National Society of Genetic Counselors (2016) does not yet support NIPT as a routine, first-tier test and recommends it for women who are considered to be at high risk for fetal aneuploidy
    • High risk patients include
      • Women ≥35 years at delivery
      • Positive biochemical serum screening test result (±NT) for T21, T18, or T13
      • Fetus with ultrasound anomalies suggesting T13, T18, T21, or MX
      • Previous pregnancy/fetus with autosomal aneuploidy
      • Parental balanced Robertsonian translocation with increased risk of T21 or T13
  • Some labs report results as positive/negative while others report a posttest risk in a format that is similar for non-DNA based aneuploidy screening tests
  • cfDNA testing is more expensive than biochemical screening tests; however, the detection and false-positive rates are greatly superior to the biochemical tests
  • Although the reported DR-SPR for cfDNA approaches that of a diagnostic test with an overall DR (including T21, T18, and T13) of 98.9% at a false-positive rate of 0.1%*, due to the rarity of some of the conditions screened, there is a real possibility of a false positive result. Thus, all positive cfDNA results should be confirmed by fetal karyotype or microarray (for microdeletions)
    • *NOTE: these values are based on the testing of high-risk women in a controlled study; detection and false-positive rates in a low-risk, uncontrolled population may be lower
  • When women have a positive cfDNA result, the positive predictive value (PPV) for the test and the abnormality should be emphasized
  • Some false positive cfDNA screens have been the result of confined placental mosaicism (CPM) or an affected vanished twin
  • If CVS is used as follow-up for confirmation of positive cfDNA results, chromosome analysis should be performed on cultured cells established from the mesenchymal core of the chorionic villi
    • Rapid aneuploidy testing by FISH or direct chromosome preparation may confirm the CPM but not reflect the fetal genotype
    • Ultimately, chromosome analysis on amniotic fluid may be required in an attempt to rule out true fetal mosaicism
  • Low fetal fraction may cause a report of “no result”
    • Low fetal fraction may be due to high maternal BMI but also is associated with increased risk for aneuploidy
  • No result due to uninterpretable cfDNA pattern is also associated with increased risk for aneuploidy; limited data is available to quantify degree of risk
  • Pergament, et al (2014) found that samples that did not yield a result (for any reason) were 2.5 times more likely to be aneuploid vs. samples that did not yield a result
  • Coexisting maternal medical conditions (eg, cancer) may affect the interpretation of cfDNA
cfDNA Specificity and Sensitivity Comparisons

Syndrome/Disorder

Method

MPSS (Palomaki, 2011 & 2012)

MPSS with SAFeR (Bianchi, 2012)

Targeted Sequencing with FORTE (Ashoor, 2012)

Panorama Test Using NATUS

Detection Rate/False-Positive Rate (%)

Trisomy 21

98.6-99.1 / 0.2

99.9 / 0.2

100 / 0.1

>99 / 0.0

Trisomy 13

91.7 / 0.9

87.5 / 0.1

80 / 0.05

>99 / 0.0

Trisomy 18

100 / 0.3

97.4 / 0.4

98 / 0.1

>99 / 0.0

Monosomy X

Not evaluated

95 / 1.0

Not evaluated

>99 / 0.0

Sex chromosome trisomies

Not evaluated

66-75%

Reported when identified

Not evaluated

>99

A NIPT/cell-free DNA screening predictive value calculator is available at https://www.perinatalquality.org/Vendors/NSGC/NIPT/ (developed by the Perinatal Quality Foundation and the National Society of Genetic Counselors [NSGC] and endorsed by ACOG and the Society for Maternal-Fetal Medicine [SMFM]

ACOG, American Congress of Obstetricians and Gynecologists; MPSS, massively parallel signature sequencing; NIPT, noninvasive prenatal testing; NSGC, National Society of Genetic Counselors; SMFM, Society for Maternal-Fetal Medicine

Recommendations, Sensitivity/Specificity, and Follow-Up

Prenatal Screening Test Information

Test

Recommended for

Purpose

First-trimester (combined) screen

Maternal screen, first trimester

Order during the first trimester (between 11w0d and 13w6d of gestation)

CRL must be between 43-83.9 mm and an NT measurement must be obtained (NT can be done when the CRL is between 38-83.9 mm)

Use when mother accepts higher SPR and wants results in the first trimester

Does not detect ONTD

Sequential screen combines first- and second-trimester screening results

First sample drawn between 11w0d and 13w6d of gestation

CRL must be between 43-83.9 mm when first-trimester blood sample is drawn and an NT measurement must be obtained (NT can be done when the CRL is between 38-83.9 mm)

Second sample drawn between 14w0d and 24w6d of gestation based on CRL

Sample 1 measures PAPP-A and total hCG

Sample 2 measures AFP, uE3, hCG, and DIA

Screens for T21, T18, ONTD

An interpretation is provided after the first draw so that pregnancies at very high risk for T21 can be identified in the first trimester

Women at intermediate or low risk after the first draw go on for the second draw and the complete screen

Integrated screen, combines first- and second-trimester screening results

First sample drawn between 10w0d and 13w6d of gestation

CRL must be between 32.4-83.9 mm when first-trimester blood sample is drawn (an NT measurement is optional for this test – must be obtained when the CRL is between 38-83.9 mm)

Second sample drawn between 14w0d and 24w6d of gestation based on CRL (if NT was provided)

Serum-only tests – do not round gestational age to nearest week; use EDD to avoid clerical errors

Sample 1 measures PAPP-A

Sample 2 measures AFP, uE3, hCG, and DIA

Screens for T21, T18, ONTD

When combined with a first-trimester certified US for NT, this test yields the best detection rate and lowest false-positive rate of all prenatal screens

Can be run without an NT (serum integrated) yielding the same detection rate with a slightly higher false-positive rate

Single Screen

(Maternal serum screen, AFP only)

Women who have had early amniocentesis, CVS, or first-trimester screening

Ideal time period is 16-18 wks of gestation; however, reference medians are available from 14w0d to 24w6d of gestation

Do not round gestational age to nearest week; use EDD to avoid clerical errors

Screen for fetal risk of ONTD at 14-25 wks

Quad Screen

Maternal serum screen, AFP, hCG, estriol, and inhibin A

ACOG recommends the quad for second-trimester aneuploidy screening for low risk women

Offer to women who

  • Present initially for second trimester
  • Do not wish to have first-trimester screening
  • Did not have access to first-trimester screening

Quad is the most economical prenatal screening test for aneuploidy

Ideal time period is 16-18 wks of gestation; however, reference medians are available from 14w0d to 24w6d of gestation

Do not round gestational age to nearest week; use EDD to avoid clerical errors

Quad screen for fetal risk of T21, T18, and ONTD

Best second-trimester screen available

Noninvasive prenatal testing (cfDNA) for fetal aneuploidy

Can be performed as early as 9.0 wks gestation

Offer to women who are considered to be at increased risk for carrying fetus with one of the common aneuploidy disorders (T21, T18, T13, or TS)

Women are considered to be at increased risk when

  • ≥35 yrs at EDD
  • Previous child with aneuploidy
  • Current fetus has US abnormalities associated with T21, T18, T13, or TS
  • Screened positive by serum screening (±NT)
  • Either parent is a carrier of a Robertsonian translocation involving chromosome 13 or 21

Highly sensitive screening test for specific fetal aneuploidies

Intended to identify women with a current pregnancy at risk for T21, T18, T13, or TS; may also identify fetuses with other sex-chromosome aneuploidies or triploidy

Women carrying a fetus with US abnormalities who screen negative by cfDNA should be offered diagnostic testing (ie, fetal karyotype and/or fetal microarray by CVS or amniocentesis)

All positive cfDNA results should be confirmed by fetal karyotype

ACOG, American Congress of Obstetricians and Gynecologists; AFP, alpha-fetoprotein; cfDNA, cell-free DNA; CRL, crown-rump length; CVS, chronic villus sampling; DIA, dimeric inhibin A; EDD, estimated due date; hCG, human chorionic gonadotropin; NT, nuchal translucency; ONTD, open neural tube defect; PAPP-A, pregnancy-associated plasma protein A; SPR, screen positive rate; uE3, unconjugated estriol; US, ultrasound
Prenatal Diagnosis: Amniotic Fluid and Chromosome Analyses
Test Recommended for Purpose

Chromosome Analysis, Chorionic Villus

Indications include

  • Increased risk for fetal aneuploidy based on maternal age, abnormal NIPT, abnormal multiple marker screening, or abnormal fetal US
  • Family history of chromosome abnormality or genetic disorder
  • Desires diagnostic testing instead of screening

Prenatal chromosome analysis on chorionic villi in pregnant woman at 10-13 wks gestation

Chorionic Villus, FISH

Rapid detection of aneuploidy involving chromosomes 13, 18, 21, X, and Y

Preliminary results usually available within 48 hrs of sample receipt by lab

Order in conjunction with fetal chromosome studies

Chromosome Analysis, Amniotic Fluid

Indications include

  • Increased risk for fetal aneuploidy based on maternal age, abnormal NIPT, abnormal multiple marker screening, or abnormal fetal US
  • Family history of chromosome abnormality or genetic disorder
  • Desires diagnostic testing instead of screening

Prenatal chromosome analysis on amniotic fluid in patient >14 wks gestation

Amniocentesis is discouraged <15 wks gestation due to high rates of fetal loss, leakage of amniotic fluid, and increased risk of club foot

Chromosome FISH, Prenatal

Rapid detection of aneuploidy involving chromosomes 13, 18, 21, X, and Y

Preliminary results usually available within 48 hrs of sample receipt by lab

Order in conjunction with fetal chromosome studies

Cytogenomic SNP Microarray – Fetal

Indications include

  • Clarification of abnormal US findings
  • Further characterize an abnormal fetal karyotype
  • Suspicion of an imbalance in a specific genomic region that is best evaluated by microarray
  • Investigate de novo, apparently balanced translocations
  • Family history of a known or suspected chromosomal abnormality best evaluated by microarray
  • Patients undergoing invasive prenatal testing (instead of, or in addition to, chromosome analysis)

Submission of a maternal blood sample for maternal cell contamination studies is encouraged

Identify genomic abnormalities (eg, aneuploidy and microdeletions) in direct or cultured amniotic fluid and CVS samples

Alpha Fetoprotein (Amniotic Fluid) with Reflex to Acetylcholinesterase and Fetal Hemoglobin

Indications include

  • Abnormal MSAFP screen
  • Family history of ONTD
  • Patient taking valproic acid or carbamazepine
  • Patient with medication-dependent diabetes or uncontrolled diabetes

Do not round gestational age to nearest week; use EDD to avoid clerical errors

Prenatal diagnosis for ONTD at 13-36 wks gestation

Amniocentesis is discouraged <15 weeks gestation due to high rates of fetal loss, leakage of amniotic fluid, and increased risk of club foot

CVS, chorionic villus sampling; EDD, estimated delivery date; MSAFP, maternal serum alpha-fetoprotein; NIPT, noninvasive prenatal testing; ONTD, open neural tube defect; US, ultrasound
Sensitivity and Initial Positive Rates for Down Syndrome

Screening test

% T21 detection

% False-positive rate

T21 cutoff

Quad

81

4-5

1/150

Integrated – serum only

85

3-4

1/110

Integrated – with NT

87

1.0

1/110

Sequential

63 (1st)
23 (2nd)
86 (total)

0.6 (1st)
1.0 (2nd)
1.6 (total)

1/25 (1st)
1/110 (2nd)

First trimester

85

6

1/230

Amniotic Fluid AChE Specificity and Sensitivity Rates for ONTD (AFP [Amniotic Fluid] with Reflex to Acetylcholinesterase)

Testing for

Sensitivity

ONDT

95%

Anencephaly

97%

Open spina bifida

99%

Abdominal wall defects

40-79%

Targeted Ultrasound Test Results Follow-Up

Test Result

Next Action

Anomaly detected on US

Perform second-tier screening

  • cfDNA for fetal aneuploidy
    • ​Choosing secondary screening by cfDNA may delay definitive diagnosis and has the potential to miss some affected fetuses

OR

Confirm with follow-up tests

  • Alpha Fetoprotein (Amniotic Fluid) with Reflex to Acetylcholinesterase and Fetal Hemoglobin
  • Chromosome Analysis, Amniotic Fluid
  • Chromosome FISH, Prenatal
  • Cytogenomic SNP Microarray - Fetal

OR

  • Chromosome Analysis, Chorionic Villus
  • Chorionic Villus, FISH
  • Cytogenomic SNP Microarray - Fetal
  • Maternal Serum Screen, Alpha Fetoprotein (Only)

No anomaly on US, but MSAFP MoMa 2.5-3.0

Repeat MSAFP (do not repeat aneuploidy screen, only the MSAFP) 2 wks after initial draw to see if AFP MoM level is increasing or decreasing; if increasing, treat patient per next row below (no anomaly on US, but MSAFP MoM >3.0)

If decreasing, probable transient maternal-fetal bleed; monitor pregnancy

  • Review or repeat level II US (including examination for signs of placental bleeding)

OR

  • Treat patient per next row below (no anomaly on US, but MSAFP MoM >3.0)

No anomaly on US, but MSAFP MoMa >3.0

Confirm with amniotic fluid tests

  • Alpha Fetoprotein (Amniotic Fluid) with Reflex to Acetylcholinesterase and Fetal Hemoglobin
  • Chromosome Analysis, Amniotic Fluid

If AF-AFP is normal, risk increases for poor pregnancy outcome (prematurity, small-for-gestational-age infant, stillbirth)

  • Offer counseling
  • Monitor pregnancy

No anomaly on US, but hCG MoMa >3.5

Increased risk for poor pregnancy outcome (preeclampsia, imminent fetal death, small-for-gestational-age infant)

  • Offer counseling
  • Monitor pregnancy

aMoM measures are multiples of the median, calculated as the value of the substance divided by the median value based on gestational age of the fetus. Adjustments to MoM values are made for maternal weight, race, number of fetuses, and maternal medication-dependent or uncontrolled diabetes.

AF-AFP, amniotic fluid alpha-fetoprotein; AFP, alpha-fetoprotein; cfDNA, cell-free DNA; FISH, fluorescence in situ hybridization; hCG, human chorionic gonadotropin; MoM, multiple of the median; MSAFP, maternal serum alpha-fetoprotein; SNP, single nucleotide polymorphisms; US, ultrasound

Second-Trimester Maternal Serum Screening Tests – Result Patterns

AFP

hCG

uE3

DIA

Pattern

L

H

L

H

Normal, overestimated gestation, and T21

H

L

H

N

Normal, underestimated gestation

L

L

L

*

Trisomy 18, fetal death

H

H

H

H

Multiple fetuses

H

N

N

N

Spina bifida, fetal-maternal hemorrhage, ventral wall defect

VH

N

L

N

Anencephaly, fetal death

VL

H

VL

N

Mole or partial mole

L, low; H, high; N, normal; VL, very low; VH, very high; *, may be high, low, or normal; not taken into account for risk calculation

Background

Down Syndrome (T21)

Epidemiology

Incidence – 1/600 births (regardless of race or geographical location)

Risk Factors

  • Risk increases with maternal age in a sigmoid fashion
    • 20s – risk for a child born with T21 is ~1:1,500
    • 30s – risk rises dramatically
    • 40s – risk levels out to ~1:100
  • ~50% of babies with T21 are born to mothers <35 years

Pathophysiology

  • Extra chromosome 21 found in all nucleated cells
  • Mosaic T21
    • Caused by an extra chromosome 21 in some, but not all, cells
    • Clinical phenotype
      • Usually milder than non-mosaic T21
      • Can vary from normal to severely affected

Clinical Presentation

  • Moderate to severe intellectual disability
  • Characteristic facial features
    • Down-slanting palpebral fissures
    • Epicanthic folds
    • Depressed nasal bridge
    • Flat mid-face
    • Low-set ears
  • Cardiac abnormalities
    • Ventricular septal defect (VSD)
    • Endocardial cushion defect
  • Hypothyroidism
  • Leukemia

ONTD

Epidemiology

Incidence – 1/900 pregnancies (varies with racial background and geographical location)

Risk Factors

  • Poorly controlled maternal diabetes mellitus
  • Family history of neural tube defects (NTD)
  • Use of certain medications in pregnancy (eg, carbamazepine, valproic acid) independent of maternal age

Clinical Presentation

  • Most common types include spina bifida (a developmental defect of the spine and overlying skin) and anencephaly (developmental failure of the brain, skull, and overlying skin)
  • Lesions of spina bifida include the following
    • Simple meningocele
    • Lipomyelomeningocele
    • Diastematomyelia
    • Myelocystocele
    • Neuritic cyst
    • Intraspinal and intrapelvic meningoceles
    • True spina bifida occulta associated with spinal dysraphism
  • Spina bifida often results in the following sequela, but clinical severity depends on several factors, especially location and size of the lesion
    • Paralysis of the lower limbs
    • Loss of bowel and bladder control
    • Cerebral ventriculomegaly requiring shunt placement
  • Anencephaly associated with limited lifespan
    • 50% of infants are stillborn
    • Remainder of newborns die within hours or days of birth

Trisomy 18 (T18)

Epidemiology

  • Incidence – 1/3,000 births
  • Survival
    • 90% stillborn
    • Most infants die within the first year of life

Pathophysiology

Extra chromosome 18 found in all nucleated cells

Clinical Presentation

  • Severe intellectual disability
  • Heart defects
  • Failure to thrive
  • Clenched fists
  • Rocker bottom feet
  • Spina bifida
  • Nonambulatory
  • Inarticulate – survivors can learn sign language
  • Risk – increases with maternal age

ARUP Laboratory Tests

Combined First- and Second-Trimester Screening Tests

First-trimester screening test for T21 (Down syndrome) and T18

Requires NT measurement performed by an ultrasonographer certified by the FMF or NTQR

Refer to Maternal Screening Sequential Specimen #2, AFP, hCG, estriol, and inhibin A test for second-trimester screening test for T21, T18, and ONTDs

Risks provided in both first and second trimesters

A screen interpreted as “normal” misses approximately 15% of ONTD cases, 10-20% of T21 cases, and 10-20% of T18 cases, depending on the test and maternal age

Second-trimester screening test for T21, T18, and ONTD

Requires a previously submitted first-trimester Maternal Screening Sequential Specimen #1, hCG, PAPP-A, NT test

Requires NT measurement performed by an ultrasonographer certified by the FMF or NTQR

Risks provided in both first and second trimesters

A screen interpreted as “normal” misses approximately 15% of ONTD cases, 10-20% of T21 cases, and 10-20% of T18 cases, depending on the test and maternal age

AFP false positives occur with multiple gestation pregnancies, underestimated gestational age

First-trimester screening test for trisomy 21 (T21, Down syndrome), trisomy 18 (T18), and ONTD

Risks determined using a combination of first- and second-trimester serum markers, with or without first-trimester NT measurement

Risks provided after testing is completed for second-trimester Maternal Serum Screening Specimen #2 AFP, hCG, Estriol, and Inhibin A test

A screen interpreted as “normal” misses approximately 15% of ONTD cases, 10-20% of T21 cases, and 10-20% of T18 cases, depending on the test and maternal age

Second-trimester screening test for T21, T18, and ONTD

Requires a previously submitted first-trimester specimen, Maternal Serum Screening Integrated Specimen #1 PAPP-A, NT test

Risks are determined after second-trimester specimen is received, using a combination of first- and second-trimester serum markers with or without first-trimester NT measurement

A screen interpreted as “normal” misses approximately 15-20% of ONTD cases, 10-20% of T21 cases, and 10-20% of T18 cases, depending on the test and maternal age

AFP false positives occur with multiple gestation pregnancies, underestimated gestational age

First-Trimester Screening Tests

First-trimester screening test for T21 and T18

Does not include AFP for ONTD screening

Requires NT measurement performed by an ultrasonographer certified by the Fetal Medicine Foundation (FMF) or Nuchal Translucency Quality Review (NTQR)

A screen interpreted as “normal” misses approximately 15-20% of ONTD cases, 10-20% of T21 cases, and 10-20% of T18 cases, depending on the test and maternal age

Second-Trimester Screening Tests

Second-trimester screening test for T21, T18, and ONTD

A screen interpreted as “normal” misses approximately 15-20% of ONTD cases, 10-20% of T21 cases, and 10-20% of T18 cases, depending on the test and maternal age

AFP false positives occur with multiple gestation pregnancies, underestimated gestational age

Second-trimester screening test for ONTD

Order this test for PREGNANT patients only; for males or nonpregnant females, refer to AFP, serum (tumor marker) test

Noninvasive Prenatal Testing (cfDNA)

First- or second-tier screening test for the most common fetal aneuploidy disorders (trisomy 13 [T13], trisomy 18 [T18], trisomy 21 [T21] [Down syndrome], Turner syndrome [TS], sex chromosome aneuploidies [XXX, XXY, XYY], and triploidy)

Testing may be offered to pregnant women from 9 weeks 0 days gestation to term

Test may also be ordered for women who have used an egg donor or for surrogate pregnancies

Test is not recommended for women carrying triplets or higher-order multiples; who have a known twin demise; who are carrying twins and used an egg donor/surrogate; or who have had an allogenic bone marrow transplant

Not recommended when the woman or her partner is a known carrier of a translocation or other chromosome rearrangement that will not result in a fetus with one of the above disorders, has a known numerical abnormality in one of the targeted chromosomes (eg, mosaic T21 or TS)

Aneuploidy for chromosomes other than 13, 18, 21, X, and Y will not be detected

Fetal mosaicism may not be detected

Low FF may occur normally in some pregnancies and can affect the ability to report a result; women with elevated BMI are at increased risk of having a low FF (may result in increased chance of a false-positive, false-negative, or no-call test result) and should be counseled accordingly

Waiting to test until the second trimester, when FF is expected to be higher than in the first trimester, may increase the chances of obtaining a result for these women

Maternal factors (eg, BMI or current cancer diagnosis) may affect FF or cfDNA analysis

First- or second-tier screening test for the most common fetal aneuploidy disorders: trisomy 13 (T13), trisomy 18 (T18), trisomy 21 (T21) (Down syndrome), Turner syndrome (TS), sex chromosome aneuploidies (XXX, XXY, XYY), and triploidy; as well as microdeletions causing 22q11.2 deletion (DiGeorge or Velocardiofacial [VCFS] syndrome), 1p36 deletion, Angelman, Prader-Willi, and cri-du-chat (5p-) syndromes

Testing may be offered to pregnant women from 9 weeks 0 days gestation to term

Test is not recommended for women who are carrying more than one fetus or have a known twin demise; patients who have used an egg donor; surrogates who have not used their own egg; or women who have had an allogenic bone marrow transplant

Aneuploidy for chromosomes other than 13, 18, 21, X, and Y will not be detected

Fetal mosaicism may not be detected

Low FF may occur normally in some pregnancies and can affect the ability to report a result; women with elevated BMI are at increased risk of having a low FF (may result in increased chance of a false positive, false negative, or no-call test result) and should be counseled accordingly

Waiting to test until the second trimester, when FF is expected to be higher than in the first trimester, may increase the chances of obtaining a result for these women

Maternal factors (eg, BMI or current cancer diagnosis) may affect FF or cfDNA analysis

All positive cfDNA results should be confirmed by diagnostic testing

Discuss positive predictive value with patient

Genetic counseling for abnormal results recommended 

First- or second-tier screening test for the most common fetal aneuploidy disorders (trisomy 13 [T13], trisomy 18 [T18], trisomy 21 [T21] [Down syndrome], Turner syndrome [TS], sex chromosome aneuploidies [XXX, XXY, XYY], and triploidy, as well as microdeletions causing 22q11.2 deletion [DiGeorge or velocardiofacial (VCFS) syndrome])

Testing may be offered to pregnant women from 9 weeks 0 days gestation to term and to women carrying monozygotic twins

Not recommended for women who are carrying dizygotic twins, triplets, or higher-order multiples; who have a known twin demise; who have used an egg donor; who are surrogates not using their own egg; or who have had an allogenic bone marrow transplant

Aneuploidy for chromosomes other than 13, 18, 21, X, and Y will not be detected

Fetal mosaicism may not be detected

Low FF may occur normally in some pregnancies and can affect the ability to report a result; women with elevated BMI are at increased risk of having a low FF (may result in increased chance of a false-positive, false-negative, or no-call test result) and should be counseled accordingly

Waiting to test until the second trimester, when FF is expected to be higher than in the first trimester, may increase the chances of obtaining a result for these women

Maternal factors (eg, BMI or current cancer diagnosis) may affect FF or cfDNA analysis

All positive cfDNA results should be confirmed by diagnostic testing

Discuss positive predictive value with patient

Genetic counseling for abnormal results recommended

Chromosome Analysis

Prenatal chromosome analysis on chorionic villi at 10-13 weeks gestation when woman is at increased risk for fetal aneuploidy based on maternal age, abnormal NIPT, abnormal multiple-marker screening, or abnormal fetal US; has a family history of chromosome abnormality or genetic disorder; or desires diagnostic testing instead of screening

Time-sensitive test

Rapid detection of aneuploidy involving chromosomes 13, 18, 21, X, and Y

Assay offered in conjunction with fetal chromosome study

Irreversible therapeutic action should not be initiated on the basis of FISH results alone

Sample type: uncultured CVS

Indications include abnormal US findings, further characterize abnormal fetal karyotype, suspicion of imbalance in specific genomic region that is best evaluated by microarray, investigate de novo, apparently balanced translocations, family history of known or suspected chromosomal abnormality best evaluated by microarray, women undergoing invasive prenatal testing (instead of, or in addition to, chromosome analysis)

Clinical sensitivity: 100% for non-mosaic aneuploidy that is detectable by karyotype

In cases with a normal karyotype, microarray studies reveal clinically relevant copy number variations (CNV) ~6% of fetuses with a structural anomaly and ~2% whose indication is advanced maternal age or positive aneuploidy screen

Will not detect base pair mutations; very small deletions/duplications; tetraploidy; balanced rearrangements, such as translocations, inversions, and balanced insertions; low-level mosaicism; imbalances of mitochondrial genome

Failure of the array to detect an imbalance at any specific locus does not exclude the diagnosis of any disorder associated with that locus

Duplications <2 MB and deletions <1 MB may not be reported if they contain only a few genes that have no compelling association with the disease

Test results are often complex, and a CNV of uncertain clinical significance may be detected

Reflex pattern: if results of aneuploidy FISH panel are normal, genomic microarray is performed; if results are abnormal, chromosome analysis is performed

Sample type: amniotic fluid

Prenatal chromosome analysis on amniotic fluid after 14 weeks gestation when woman is at increased risk for fetal aneuploidy based on maternal age, abnormal noninvasive prenatal testing (NIPT), abnormal multiple-marker screening, or abnormal fetal US; has a family history of chromosome abnormality or genetic disorder; or desires diagnostic testing instead of screening

Time-sensitive test

Genetic counseling for abnormal results

Sample type: amniotic fluid

Indications include abnormal US findings, abnormal prenatal screen, fetal demise

Clinical sensitivity: 100% for non-mosaic aneuploidy that is detectable by karyotype

Does not detect base pair mutations; very small deletions/duplications; tetraploidy; balanced rearrangements, such as translocations, inversions, and balanced insertions; low-level mosaicism; imbalances of mitochondrial genome

Failure of the array to detect an imbalance at any specific locus does not exclude the diagnosis of any disorder associated with that locus

Duplications <2 MB and deletions <1 MB may not be reported if they contain only a few genes that have no compelling association with the disease

Test results are often complex, and a CNV of uncertain clinical significance may be detected

Reflex pattern: if results of chromosome analysis are normal, genomic microarray is performed; in cases with a normal karyotype, microarray studies reveal clinically relevant copy number variations (CNV) in ~6% of fetuses with an US anomaly and ~2% whose indication is advanced maternal age or positive aneuploidy screen

Sample type: uncultured amniotic fluid

Rapid detection of aneuploidy involving chromosomes 13, 18, 21, X, and Y

Assay offered in conjunction with fetal chromosome study 

Does NOT detect structural chromosome abnormalities, mosaicism, and other numerical chromosome abnormalities (excluding 13, 18, 21, X, and Y)

Genetic counseling for abnormal results

Irreversible therapeutic action should not be initiated on the basis of FISH results alone

Sample type: uncultured amniotic fluid

For rapid detection of aneuploidy involving chromosome 13, 18, 21, X, and Y

In cases with a normal karyotype, microarray studies reveal clinically relevant copy number variations (CNV) ~6% of fetuses with a structural anomaly and ~2% whose indication is advanced maternal age or positive aneuploidy screen

Will not detect base pair mutations; very small deletions/duplications; tetraploidy; balanced rearrangements, such as translocations, inversions, and balanced insertions; low-level mosaicism; imbalances of mitochondrial genome

Failure of the array to detect an imbalance at any specific locus does not exclude the diagnosis of any disorder associated with that locus

Duplications <2 MB and deletions <1 MB may not be reported if they contain only a few genes that have no compelling association with the disease

Test results are often complex, and a CNV of uncertain clinical significance may be detected

Irreversible therapeutic action should not be initiated on the basis of FISH results alone

Reflex pattern: if results of aneuploidy FISH panel are normal, genomic microarray is performed; if results are abnormal, chromosome analysis is performed

Sample type: direct or cultured amniotic fluid and CVS samples

Diagnostic test designed to identify genomic abnormalities (eg, aneuploidy and microdeletions)

Indications include clarification of abnormal US findings, further characterize an abnormal fetal karyotype, suspicion of an imbalance in a specific genomic region that is best evaluated by microarray, investigate de novo, apparently balanced translocations, family history of a known or suspected chromosomal abnormality that is best evaluated by microarray, patients undergoing invasive prenatal testing (instead of, or in addition to, chromosome analysis)

Submission of a maternal blood sample for maternal cell contamination is encouraged

Clinical sensitivity: 100% for non-mosaic aneuploidy that is detectable by karyotype

In cases with a normal karyotype, microarray studies reveal clinically relevant copy number variations (CNV) ~6% of fetuses with a structural anomaly and ~2% whose indication is advanced maternal age or positive aneuploidy screen

Time-sensitive test

Does not detect base pair mutations; very small deletions/duplications; tetraploidy; balanced rearrangements (translocations, inversions, and balanced insertions); low-level mosaicism; imbalances of the mitochondrial genome

To minimize variations of undetermined significance, duplications <2 MB and deletions <1 MB may not be reported if they contain only a few genes that have no compelling association with disease

May reveal a biological relationship, including a close blood relationship (consanguinity or incest) between the mother and the father of the fetus

Failure of the array to detect an imbalance at any specific locus does not exclude the diagnosis of any disorder associated with that locus

Genetic counseling for abnormal results

Related Tests

Aid in the diagnosis and monitoring of various hormonal reproductive disorders

Not a prenatal test

Use following an abnormal amniotic fluid AFP result to evaluate possibility of a fetal ONTD

Evaluate possibility of a fetal ONTD at 13-36 weeks of gestation

Prenatal testing for fetus with ultrasound findings suggestive of Noonan syndrome, such as cystic hygroma, increased NT, or polyhydramnios

Acceptable initial test to confirm a clinical diagnosis of NS or LEOPARD syndrome

Clinical sensitivity: ~50-60% for NS and 90% for LEOPARD syndrome

Panel is ONLY utilized for newborns with suspected aneuploidy

Panel includes probes for trisomy 13, trisomy 18, trisomy 21, and chromosomes X and Y

If chromosome analysis is also ordered, a preliminary chromosome report will be issued based on 10 cells from a 48-hour culture

Medical Experts

Contributor

Baldwin

Erin Baldwin, MS, LCGC
Manager, Genetic Counselor Services, ARUP Laboratories
Contributor

Genzen

Jonathan R. Genzen, MD, PhD
Associate Professor of Clinical Pathology, University of Utah
Chief Operations Officer: Medical Director, Automated Core Laboratory, ARUP Laboratories
Contributor
Contributor

Lamb

Allen N. Lamb, PhD, FACMG
Allen N. Lamb, PhD, FACMG
Retired Former Professor of Clinical Pathology, University of Utah
Retired Former Laboratory Section Chief, Cytogenetics and Genomic Microarray, ARUP Laboratories
Contributor

Mao

Rong Mao, MD, FACMG
Professor of Clinical Pathology, University of Utah
Section Chief, Molecular Genetics and Genomics, ARUP Laboratories

References

Additional Resources
Resources from the ARUP Institute for Clinical and Experimental Pathology®