PROFESSIONAL DEVELOPMENT PAPER

NSGC Practice Guideline: Prenatal Screening and Diagnostic

Testing Options for Chromosome Aneuploidy

K. L. Wilson & J. L. Czer winski & J. M. Hoskovec &

S. J. Noblin & C. M. Sullivan & A. Harbison &

M. W. Campion & K. Devary & P. Devers &

C. N. Singletary

Received: 1 June 2012 / Accepted: 17 September 2012 / Published online: 22 November 2012

National Society of Genetic Counselors, Inc. 2012

Abstract The BUN and FASTER studies, two prospec-

tive mult icenter trials in the United States, validated the

accuracy and detection rates of first and second trimester

screening previously reported abroad. These studies, cou-

pled with the 2007 release of the American College of

Obstetricians and Gynecologists (ACOG) Practice Bulle-

tin that endorsed first trimester screening as an alternative

to traditional s econd trimester multiple m arker screening,

led to an explosion of screening options available to

pregnant women. ACOG also recommended that invasive

diagnostic testing for chromosome aneuploidy be made

available to all women regardless of maternal age. More

recently, another option known as Non-invasive Prenatal

Testing (NIPT) became available to screen for chromo-

some aneuploidy. While screening a nd test ing opt ions

may be lim ited due to a variety of fact ors, healthcare

providers need to be aware of the options in their area

in order to provide their patients with accurate and reli-

able information. If not presented clearly, patients may

feel overwhelmed at the number of choices available. The

following guideline includes recommendations f or health-

care providers regarding which sc reening or diagnostic

test should be offered based on availability, insurance

coverage, a nd ti ming o f a patient’s entry into prenatal

care, as well as a triage assessment so that a ge neral

process can be adapted to unique situations.

Keywords Prenatal screening

Prenatal testing

Chromosome aneuploidy

Genetic counseling

National

Society of Genetic Counselors

Practice guidelines

Purpose

To provide information that assists physicians and allied health

professionals in making decision s about diff erent screening and

diagnostic testing for chromosome aneuploidy throughout

pregnancy.

K. L. Wilson (*)

Department of Ob/Gyn, Division of Gynecologic Oncology,

The University of Texas Health Science Center at Houston,

6410 Fannin St, Suite 1217,

Houston, TX 77030, USA

e-mail: [email protected]

S. J. Noblin

C. N. Singletary

Department of Pediatrics,

The University of Texas Health Science Center at Houston,

6410 Fannin Street, Suite 1217,

Houston, TX 77030, USA

P. Devers

Department of Obstetrics and Gynecology,

Division of Maternal-Fetal Medicine,

University of North Carolina at Chapel Hill,

CB# 7516,

Chapel Hill, NC 27599-7516, USA

A. Harbison

Deparment of Ob/Gyn, Division of Maternal

Fetal Medicine, The University of Texas

Health Science Center at Houston,

5656 Kelley Street,

Houston, TX 77026, USA

M. W. Campion

Boston University School of Medicine,

85 E. Newton St, M913,

Boson, MA 02118, USA

K. Devary

EvergreenHealth Maternal Fetal Medicine,

12333 NE 130th Lane, Suite Tan 240,

Kirkland, WA 98034, USA

J. L. Czerwinski

J. M. Hoskovec

C. M. Sullivan

Department of Ob/Gyn, Division of Maternal Fetal Medicine,

The University of Texas Health Science Center at Houston,

6410 Fannin St, Suite 1217,

Houston, TX 77030, USA

J Genet Counsel (2013) 22:4–15

DOI 10.1007/s10897-012-9545-3

Disclaimer

The practice guidelines of the National Society of Genetic

Counselors (NSGC) are developed by members of the NSGC

to assist genetic counselors and other health care providers in

making decisions about appropriate management of genetic

concerns; including access to and/or delivery of services. Each

practice guideline focuses on a clinical or practice-based issue,

and is the result of a review and analysis of current profes-

sional literature believed to be reliable. As such, information

and recommendations within the NSGC practice guidelines

reflect the current scientific and clinical knowledge at the time

of publication, are only current as of their publication date,

and subject to change without notice as advances emerge.

In addition, variations in practice, which take into account

the needs of the individual patient and the resources and

limitations unique to the institution or type of practice, may

warrant approaches, treatments, and/or procedures that differ

from the recommendations outlined in this guideline. There-

fore, these recommendations should not be construed as dic-

tating an exclusive course of management, nor does the use of

such recommendations guarantee a particular outcome. Ge-

netic counseling practice guidelines are never intended to

displace a health care provider’s best medical judgment based

on the clinical circumstances of a particular patient or patient

population. Practice guidelines are published by NSGC for

education and informational purposes only, and NSGC does

not “approve” or “endorse” any specific methods, practices, or

sources of information.

Background

The landscape of prenatal screening changed in 2007 with

the release of two ACOG Practice Bull etins stating all

women should be offered maternal serum screening (MSS)

and diagnostic testing regardless of maternal age, and that

healthcare provide rs should determine which screening

options would best serve their patients (ACOG Practice

Bulletin No. 77 & 88, 2007). ACOG’s assertion was subse-

quently echoed by the American College of Medical Genet-

ics (ACMG), leaving providers to reassess their screening

and diagnostic testing practices (Driscoll and Gro ss 2008).

Prenatal screening strategies have evolved greatly over the

years. Various combinations of firs t and second trimester

maternal serum analytes and fetal ultrasound findings have

been proposed as part of an ongoing quest to create a screen-

ing test with the highest detection and lowest false positive

rates. Recently, NIPT has been made commercially available

as an alternative option for chromosome aneuploidy screen-

ing. Screening options for chromosome aneuploidy are non-

invasive, which may make them attractive options for patients

who desired more individualized risk assessment information

prior to making a decision about whether or not to undergo

invasive diagnostic testing. The primary limitation of screen-

ing is that it does not provide a definitive diagnosis, leading to

the potential of increased anxiety in women with an unaffect-

ed pregnancy and the potential of false reassurance in women

who have a pregnancy with a chromosome aneuploidy. An-

other limitation of screening is the variability in the detection

rates, false positive rates, screening cut-offs, and anatomical

ultrasound markers included in the screen based on the par-

ticular laboratory and/or provider involved. In addition, detec-

tion rates for screening in multiple gestations are generally

decreased from those of singletons (Wald and Rish 2005). For

a more comprehensive review of the advantages and limita-

tions of the types of screening, refer to the individual sections

below and to Tables 1, 2,and3. A decision tree to assist

providers in selecting a screening method that is most suitable

for their practice is presented in Fig. 1.

In addition to the various prenatal screening options, diag-

nostic testing for chromosomal abnormalities (Table 4)is

available. Chorionic villus sampling, or CVS, is typically

performed between 10-13w6d of gestation, and involves a

transcervical or transabdominal aspiration of chorionic villi

from the developing placenta (Wapner 2005). Another option

for diagnostic testing is amniocentesis. Amniocentesis is tra-

ditionally performed after 15 weeks of gestation by trans-

abdominal removal of amniotic fluid (CDC 1995). Samples

obtained through CVS and amniocentesis are typically used

for chromosomal analysis by karyotyping, but may also be

used for rapid interphase fluorescence in situ hybridization

(FISH) to screen for chromosome aneuploidy, metaphase

FISH to evaluate for specific microdeletions or microduplica-

tions, chromosomal microarray, or other molecular testing.

A referral for genetic counseling has traditionally been

made for patients who have an increased risk for chromosome

aneuploidy, including those with a positive maternal serum

screen result, positive family history, a fetal anomaly identi-

fied on ultrasound, or those who are 35 years of age or older at

delivery. However, as screening options have expanded, it has

become more routine to refer patients for genetic counseling

when they are having difficulty deciding on a course of action

for screening or diagnostic testing. Appropriate screening and

diagnostic testing options are typically presented to patients in

the context of a prenatal genetic counseling session (CDC

1995). Genetic counselors are uniquely trained to explain

complex information in an understandable format to patients

and to facilitate the informed decision-making process.

First Trimester Screening Options

The first trimester analyt es pregnancy associa ted plasma

protein-A (PAPP-A) and free beta-human chorionic gonad-

otropin (β-hCG) may be measured between 9w0d-13w6d of

gestation, while the nuchal translucency (NT) measurement

NSGC Practice Guideline 5

Table 1 Screening options for chromosome aneuploidy in pregnancy

First

Trimester

Analyte

Combined

First

Trimester

Integrated Serum Integrated Stepwise

Sequential

Contingency Multiple

Marker

Serum

Screening

Non Invasive

Prenat al

Testing

First

visit

Second

visit

First visit Second

visit

First visit Second

visit

First visit Second

visit

Gestational

Age at

Blood Draw

9w0d-

13w6d

9w0d-

13w6d

9w0d-

13w6d

15w0d-

21w6d

9w0d-

13w6d

15w0d-

21w6d

9w0d-

13w6d

15w0d-

21w6d

9w0d-

13w6d

15w0d-

21w6d

15w0d-

21w6d

10w0d -

21w6d

Maternal

Serum

Analytes

PAPP-A

b-hCG

(or hCG)

PAPP-A

b-hCG

(or hCG)

PAPP-A AFP

hCG

uE3

DIA

PAPP-A AFP

hCG

uE3

DIA

PAPP-A b-hCG

(or hCG)

AFP

hCG

uE3

DIA

PAPP-A b-hCG

(or hCG)

AFP

hCG

uE3

DIA

AFP

hCG

uE3

DIA

ITA

Circulating

Cell Free

Fetal DNA

NT Utilized No Yes

Yes

No Yes

Yes

No No

Down Syndrome

Detection Rate

62–63 %

78–91 %

n/a

94–96 %

n/a

87–88 %

91–95 %

91–92 %

75–83 %

99–100 %

11, 12

Trisomy 18

Detection Rate

82 %

91–96 %

n/a

91–96 %

n/a

82 %

91–96 %

60–70 %

97–100 %

11, 12

Provides

ONTD Risk

No No n/a

Yes n/a

Yes No Yes

No Yes

Yes No

•Sources: Barkai et al. (1993); Bianchi et al. (2012); Cole et al. (1999); Cuckle et al. (2008); Haddow et al. (1998); Malone et al.(2005); Norton et al. (2012); Palomaki et al. (2006); Palomaki et al.

(2012); Spencer and Nicolaides (2002); Wald et al. (2003); Wald et al. (2004); Wapner et al. (2003)

1. Whole molecule hCG used in place of free b-hCG by some laboratories and some laboratories add additional analytes such as DIA, both of which may change detection rates

2. Typically at 5 % false positive rate (FPR) with 1/270 cut-off for screen positive; if other FPR or cut-off used, may change detection rate; FPR averages 15–20 % if AMA

3. Typically at 0.5 % FPR with 1 in 100 cut-off, if other FPR or cut-off used, may change detection rate; some laboratories quote combined t18/t13 risk & detection rate

4. The timing of assessment for the NT measurement is between 10w4d and 13w6d

5. Results not provided after first visit

6. Second blood draw is required for all patients

7. Second blood draw is indicated for a portion of patients after the first step of the screen, but not all patients

8. NTD risk assessment is performed for those patients who need a second step and thus get a blood draw that contains AFP as an analyte

9. ITA is included by one laboratory along with AFP, hCG, uE3, and DIA and referred to as the Penta Screen; detection rates are similar to the Quadruple Screen

10. Cell free fetal DNA is found in maternal blood but is not considered an analyte like traditional serum screening

11. While initial studies show a range of detection rates up to 100 % for trisomy 18 & 21, NIPT is not yet considered diagnostic and follow-up CVS or amnio is recommended for a positive NIPT

12. Typically <1 % FPR (from 0.1 to 0.97 %); laboratories generally report trisomy 13 as well, with detection ranging 79-92% at <1 % FPR.

6 Wilson et al.

is valid when measured between 10w4d-13w6d of gestation.

It is important to note that some laboratories substitute hCG

for β-hCG, which may impact detection rates. Most labora-

tories target a 5 % false positive rate for serum screening

options.

There are three options for first trimest er screening. NT

only with maternal age relies upon the size of the fetal NT

and the patient’s age to calculate a risk estimate. First

trimester analyte screening relies upon the levels of mater-

nal serum PAPP-A and free β-hCG or total hCG combined

Table 2 Aneuploidy screening options comparison table

Advantages Limitations Physician likely to utilize screen

First

Trimester

Analyte

Screening

• 1st trimester result

• NT not required

• 1 visit

• CVS is an option if screen positive

• Lower detection rate compared to other

screening options that include NT

Physician with an early-to-care patient

population who does not have access

to a certified NT provider, but does

have access to CVS, and prefers to

complete screening in one visit.

Combined

First

Trimester

Screening

• 1st trimester result

• 1 visit

• CVS is an option if screen positive

• NT required

• Lower detection rate compared to integrated

screen

Physician with an early-to-care patient

population who has access to certified

NT provider and to CVS, and prefers

to complete screening in one visit.

Integrated

Screening

• Highest detection rates out of all of

maternal serum screening tests

• 2 visits

• NT required

• Results given in 2nd trimester

Physician with an early-to-care patient

population who has access to certified

NT provider, but does not have access

to CVS.

Serum

Integrated

Screening

• Highest detection rates for screening

when the NT is not available

• NT not required

• 2 visits

• Results given in 2nd trimester

• Lower detection rate compared to screens that

include NT, such as integrated screen

Physician with an early-to-care patient

population who does not have access

to certified NT provider or to CVS.

Stepwise

Sequential

Screening

• 1st trimester result for highest risk

patients allows option of CVS

• Detection rate higher than combined

FTS while still allowing for some 1st

trimester results

• 2 visits for most patients

• NT required

• Lower detection rate compared to integrated

screen

Physician with an early-to-care popu-

lation and high follow-up compliance

who has access to a certified NT pro-

vider and to CVS, who wants infor-

mation early enough to offer CVS if

risk is high, and wants to avoid mod-

erate risk group created by contingency

screening.

Contingency

Screening

• 1st trimester results for high and low

patients, minimizing number of patients

needing a second visit

• 2 visits for moderate risk group

• NT required

•

Initial moderate risk group may not feel as

reassured with 2nd trimester negative screen

results as initial low risk group

Physician with an early-to-care patient

population and high follow-up com-

pliance who has access to a certified

NT provider and to CVS, and who

feels the benefit of a one visit screen

for most patients outweighs anxiety

caused for patients who fall into the

moderate risk group and are later re-

stratified to a low-risk group.

Multiple

Marker

Serum

Screening

• Allows for screening in women

presenting for care after the first

trimester

• 1 visit

• Results given in 2nd trimester

• Lower detection rate compared to screens that

include first trimester components and/or NT

Physician who has patients who

present primarily in second trimester

for screening or patients whose

insurance does not cover NT screening.

Non Invasive

Prenatal

Testing

• 1st or 2nd trimester result in 1 visit

without NT required

• Highest detection rates across all non

invasive options

• New technology with shorter publication

history and less information on payer coverage

Physician comfortable with new

technology who wants a screening test

with high detection rate and low false

positive rate that can be applied in

both first and second trimester.

NSGC Practice Guideline 7

with maternal age for risk estimation. The third method is

referred to as combined first trimester screening, because it

combines the use of the NT meas urement, the first trimester

maternal serum analytes (PAPP-A, and free β-hCG or total

hCG) and maternal age. Combined first trimest er screening

has been demonstrated to have higher detection rates for

Down syndrome (78–91 %) and trisomy 18 (91–96 %)

compared to NT only or serum analyte only m etho ds

(Malone et al. 2005;Waldetal.2003;Wapneretal.

2003), (Refer to Tables 1 and 3). Some studies have

reported that pregnancies affected with trisomy 13 have

an analyte and NT pattern sim ilar to tri somy 18; thus, it

may also be possible to use combined first trimester

screening to screen for trisomy 13 (Spencer et al. 2000).

When the NT measurement is significantly elevated (typi-

callyconsideredtobeanNT≥3.0 mm or the 95th percentile),

the possibility of chromosome aneuploidy is significantly

increased, and it is appropriate to offer the option of diagnostic

testing at that point without performing or waiting for the

results of the MSS (Comstock et al. 2006). In addition to

chromosome aneuploidies, NT ≥3.5 mm is also associated

with an increased risk of a congenital heart defect. In such

cases, fetal echocardiography is recommended during the

second trimester. It is important to note that once an elevated

NT is seen, the risk for an adverse pregnancy outcome remains

increased even if normal fetal karyotype is confirmed (Bilardo

et al. 2010).

As none of the first trimester screening options allow for

a calculation of the risk for open neural tube defects

(ONTD), a maternal serum alpha-fetoprotein (MSAFP)

sample should be drawn in the second trimester in patients

undergoing first trimester screening. ONTDs may also be

detected on second trimester ultrasound. It is not recom-

mended that an independent risk assessment for chromo-

some aneuploidy be performed in both the first and second

trimesters due to the high false positive rate (Platt et al.

2004).

The presence or absence of the fetal nasal bone and of

tricuspid regurgitation on first trimester ultrasound have

been proposed as additional ultrasound markers for chromo-

some aneuploidy (Kagan et al. 2009 ; Ozkaya et al. 2010).

However, at the present time, the use of these anatomical

markers to determine chromosome aneuploidy risk is not

universally recommended.

Non-Invasive Prenatal Testing (NIPT) NIPT uses circulat-

ing cell free fetal DNA in maternal plasma to evaluate for

Down syndrome, trisomy 18, and trisomy 13. Published

studies show a low false posit ive rate (<1 %) and a very

high detection rate for Down syndrome (99–100 %), triso-

my 18 (97–100 %) and trisomy 13 (79 – 92 %), (Bianchi et

al. 2012; Norton et al. 2012; Palomaki et al. 2012), (see

Table 1). In addition, Bianchi et al. fo und a 94 %

detection rate for monosomy X and reported several

Table 3 Ultrasound screening options for aneuploidy

Nuchal Translucency FetalAnatomy Ultrasound

Gestational Age 10w4d–13w6d 18w0d–20w6d (ideal)

16w0d–24w6d (at some centers)

Ultrasound Markers

Evaluated

Increased Nuchal

Translucency

1,2

Structural Defects, e.g., congenital heart defects, hydrocephalus, holoprosencephaly, cystic

hygroma, cleft lip, duodenal atresia, omphalocele, kidney malformations, intrauterine growth

restriction, clenched hands, etc.

Soft Markers, e.g., increased nuchal fold, choroid plexus cyst, echogenic bowel, hydronephrosis,

echogenic focus/foci, single umbilical artery, clinodactyly of the 5th finger, shortened long bones

Down Syndrome

Detection Rate

60–82 %

3,4

50–70 %

4,5

Trisomy 18

Detection Rate

71–82 %

3,4

80%

4,5

Provides ONTD

Risk Assessment

No Yes

•Sources: ACOG, 2009; Malone et al. 2005; Nicolaides et al. 2002; Nyberg and Souter 2001; Sanders et al. (2002); Taslimi et al. 2005; Wald et al.

2003; Wapner et al. 2003

1. Defined as>95th percentile or >3.0 mm by the majority of studies

2. Other ultrasound markers such as nasal bone and tricuspid regurgitation remain early in investigation and are not included

3. False positive rate 20–25 %; includes age in detection rate; without age detection is lower

4. Detection rate is affected by gestational age, quality of ultrasound images, operator experience, equipment used, referral indication, criteria for

positive findings, maternal habitus and fetal position

5. Adjustment of a priori risk based on the association between a given ultrasound finding and aneuploidy varies by center. The magnitude of

increased risk is impacted by the specific findings and the strength of their association with aneuploidy

8 Wilson et al.

ca ses of mosaicism for trisomy 21, trisomy 18, and mono-

somy X (2012). The landmark NIPT studies recruited

women who were otherwise pursuing i nvasive testing.

Thus, the major ity were from high-risk populations that

had an increased risk to have a fetus affected with chromo-

some aneuploidy. At this time, NIPT is only recommended for

patients from high-risk populations, including advanced ma-

ternal age, positive screening test, abnormal ultrasound sug-

gestive of aneuploidy, or prior pregnancy with chromosome

aneuploidy (Devers et al. 2012). It is recomme nded t hat a

positive NIPT be followed by confirmatory diagnostic

testi ng prior t o making pregnancy decisions (Benn et al.

2011; Devers e t al. 2012). NIPT is validated between

10w0d–21w6d gestation, making it an option for wom-

en that present in either the first or the second trimester.

Many practices are using NIPT in addition to their

already established screening practices to provide high-

risk patients with m ore information prior to m aking a

decision about invasive diagnostic testing. If NIPT is

performed, additional serum screening for chromosome

aneuploidy is not recommended. While NIPT has prom-

ise for the future and may potentially replace other

screening methods as the standard of care, there is s till

much to learn about this technology and its clinical

utility. For more information regarding N IPT, please refer

to the fact sheet published in 2012 by the National

Coalition for Health Professional Education in Genet ics

and NSGC.

Integrated, Serum Integrated, Stepwise Sequential

and Contingency Screening

Screening strategies that combine both first and second

trimester serum analytes and ultrasound markers have been

proposed as a means of increasing detection rates while

decreasing false positive rates. Since these screening options

involve multiple steps, it may be difficult to ensure that all

Following appropriate counseling, does patient

wish to pursue prenatal screening or testing?

YES

Patient elects

diagnostic testing

Chorionic

Villus

Sampling

between

10 and 13

weeks

Amnio -

centesis

>15 weeks

Patient elects screening

prior

to making a decision about

diagnostic testing

Patient fits current

criteria for NIPT &

elects NIPT

NIPT

or 2

trimester

result

1 visit

Patient elects maternal

serum screening

< 14

weeks

NOT

available/feasible

First

Trimester

Analyte

trimester

result

1 visit

Serum

Integrated

trimester

result

2 visits

available/feasible

Combined

FTS

trimester

result

1 visit

Stepwise

Sequential

trimester

result if very

high risk

trimester

result if less

risk

1-2 visits

Integrated

trimester

result

2 visits

Greatest

analyte

detection

rate

Contingency

trimester

result if high or

low risk

trimester

result for

moderate risk

1-2 visits

> 14

weeks

Quadruple

Marker

Screening

1 visit

See Table 4 for more information about diagnostic testing.

See Table 1 for more information about screening options.

NT= Nuchal translucenc

Fig. 1 Decision tree for selecting between screening and diagnostic testing options

NSGC Practice Guideline 9

patients return for the multiple visits required to complete

the screen ing process (Wald et al. 2003).

Integrated Screening uses a two-step process to adjust the

maternal age-related risk for Down syndrome and trisomy

18. The first step involves NT and PAPP-A measurements in

the first trimester, and the second step involves serum analyte

measurements of AFP, hCG, unconjugated estriol (uE3), and

dimeric inhibin A (DIA) in the second trimester (Cuckle et al.

2008;Maloneetal.2005;Waldetal.2003). The patient is

given a single risk assessment after both steps have been

completed. Integrated screening has a high detection rate for

Down syndrome (94–96 %) and trisomy 18 (91–96 %); how-

ever , patients must wait until the second trimester for results,

which eliminates the opportunity for early diagnostic testing

such as CVS (Malone et al. 2005; Spencer and Nicolaides

2002;Waldetal.2004;Wapneretal.2003). When an NT

measurement is not possible, serum integrated screening may

still be performed at a lower detection rate for Down syndrome

(87–88 %) and trisomy 18 (82 %), (Malone et al. 2005;Wald

and Rish 2005;Wapneretal.2003).

Stepwise Sequential Screening also involves two steps,

with the first step combining the NT measurement, serum

analytes PAPP-A and β-hCG (or hCG), and maternal age in

the first trimester. Patients are initially stratified into a

“high” or “low” risk group as determined by the screening

laboratory’s risk cut-off figures. These initial results are

disclosed to patients who fall into the high risk group, and

they are subsequently offered diagnostic testing. Initial results

may or may not be disclosed to patients who fall into the low

risk group, and they proceed with a second blood draw per-

formed in the second trimester. In this second step, the serum

analytes AFP, hCG, uE3, and DIA are incorporated into the

risk assessment, and the patient is given a risk for chromo-

some aneuploidy based on the combined results of both of the

screening steps. The overall detection rate for stepwise se-

quential screening is 91–95 % for Down syndrome and 91–

96 % for trisomy 18 (Malone et al. 2005;Palomakietal.2006;

Spencer and Nicolaides 2002; Wapner et al. 2003). Stepwise

sequential screening allows the patients with the highest risk

to consider diagnostic testing in the first trimester of pregnan-

cy while retaining some of the increase in detection rates seen

with integrated screening.

Contingency Screening is another two-step screening option.

As with stepwise sequential screening, the first step adjusts

maternal age-related risk for chromosome aneuploidy based

on the NT measurement and serum analytes. However , after

the first step, patients are divided into low, moderate, and high

risk groups based on their risk assessment for chromosome

abnormality. The high risk group is offered diagnostic testing,

the moderate risk group continues to the second serum analyte

screening step, and the low risk group is not offered further

serum analyte screening. Once the moderate risk group under-

goes second trimester analyte screening (AFP, hCG, uE3, and

Table 4 Diagnostic testing options

Chorionic Villus Sampling (CVS) Early Amniocentesis

Amniocentesis

Gestational Age at

Time of Procedure

10w0d–13w6d 11w0d–13w0d 15w0d–23w6d

Test Methodology Transcervical or transabdominal

aspiration of chorionic villi from

developing placenta

Abdominal withdrawal of amniotic

fluid from gestational sac

Risk of Miscarriage 0.5–1.0 % 2.0 % 0.2–0.3 %

Additional Associated Risks

• Mosaicism: <1 % • Mosaicism: 0.2 % • Mosaicism: 0.2 %

• Maternal Cell Contamination: <1 % • Club foot: 1.3% –1.7 % • Club foot:<0.1 %

• Spotting & Cramping: 15 % • Amniotic Fluid Leakage:

3.5 % – 4.4 %

• Amniotic Fluid Leakage: 1.7 %

Down Syndrome Detection Rate

98–99 % 99.80 % 99.90 %

Trisomy 18 Detection Rate

98–99 % 99.80 % 99.90 %

Provides ONTD Detection No Yes Yes

Sources: ACOG Practice Bulletin No. 88, 2007; Brambati et al. 1992; Canadian Collaborative CVS Amniocentesis Clinical Trial Group, 1989;

Centers for Disease Control and Prevention Morbidity and Mortality Weekly Report, 1995; Eddleman et al. 2006; Jackson et al. 1992; Ledbetter et

al. 1992; Sundberg et al. 1997; The Canadian Early and Mid-trimester Amniocentesis Trial Group, 1998; Wapner et al. 2003; Winsor et al. 1999

1. May be performed at other gestational ages, but risks and benefits may vary

2. An increased risk for limb reduction defects has been associated with CVS performed prior to 10 weeks gestation

3. Detection rates based on karyotyping; other detection rates may apply when utilizing fluorescence in situ hybridization or chromosomal

microarray

4. Early amniocentesis is not recommended by ACOG

5. Range of 0.06–1.0 % [or 1 in 1600 to 1 in 100] reported in literature

10 Wilson et al.

DIA), these patients are then stratified again into a high risk

group who is offered diagnostic testing and a low risk group

who is not considered at great enough risk to warrant diagnostic

testing. The overall detection rate for contingency screening

when both steps are performed is 91–92 % for Down syndrome

and 91–96 % for trisomy 18 (Cuckle et al. 2008;Palomakietal.

2006; Spencer & Nicolaides 2002;Wapneretal.2003). Con-

tingency screening lowers the number of patients proceeding to

the second trimester serum analyte screening step compared to

stepwise sequential screening, while still identifying the major-

ity of high risk pregnancies in the first trimester. However,

patients identified as initially having a moderate risk may retain

some anxiety even if the second step places them back into the

low risk group.

Second Trimester Screening Options

Second trimester MSS options include the triple screen, the

quadruple (quad) screen, and the penta screen. The triple

screen consists of maternal serum AFP, hCG, and uE3 com-

binedwithanaprioririskbasedonmaternalagetoscreenfor

Down syndrome, trisomy 18, and ONTDs. The quad screen

adds DIA, which increases the detection rate for Down syn-

drome (Benn et al. 2003). One laboratory has added invasive

trophoblast antigen (ITA) to the quad screen and named it the

penta screen, although detection rates are not substantially

different from the quad screen (Cole et al. 1999). Second

trimester maternal serum screens are typically performed be-

tween 15w0d-21w6d gestation (Wald et al. 2004). Detection

rates for Down syndrome (75–83 %) and trisomy 18 (60–70 %)

are highest with the quad screen or penta screen (Table 1),

(Wald et al. 2003). Detection rates for ONTDs are the same

for all second trimester screening options (Barkai et al. 1993;

Haddow et al. 1998;Waldetal.2004). Second trimester screens

are helpful for women who first present for prenatal care in their

second trimester, but all second trimester MSS have lower

detection rates compared to screening options that incorporate

first trimester serum analytes. NIPT may represent a viable

alternative for screening with a high detection rate for eligible

patients who present in the second trimester.

A fetal ultrasound in the second trimester may also be used

to screen for chromosome aneuploidy by identifying both true

structural defects and structural variants, which are commonly

referred to as “soft markers” (Table 3). Ultrasound may also

detect isolated structural defects not typically associated with

chromosome aneuploidy. The optimal time to perform an

ultrasou nd to survey fetal anatomy is between 18-20w6d

gestation (ACOG Practice Bulletin No. 101, 2009). The risk

for chromosome aneuploidy is dependent upon not only the

combination of soft markers and/or structural defects seen, but

also the expertise of the provider interpreting the ultrasound

(Nyberg and Souter 2001; Sanders et al. 2002). Chromosome

aneuploidy screening by ultrasound can further be limited by

maternal habitus and fetal position. Approximately 50–70 %

of pregnancies with Down syndrome and 80 % of pregnancies

with trisomy 18 will have anatomical defects and/or markers

identified by the detailed or targeted ultrasound (Nyberg and

Souter 2001). Based on these detection rates, second trimester

anatomy ultrasound alone is not as accurate as MSS or NIPT.

Other ultrasounds, such as growth scans, may also detect

structural anomalies that are associated with chromosome

aneuploidy, but these ultrasounds are not routinely used for

chromosome aneuploidy screening.

Diagnostic Testing Options

Current procedures available for the diagnosis of chromo-

some aneuploidy during pregnancy include: CVS, early

amniocentesis, and amniocentesis (Table 4).

Chorionic Villus Sampling (CVS)

CVS is used for the detection of fetal chromosome aneuploidy

in the first trimester. The accuracy for detection of fetal chro-

mosome abnormalities by CVS is estimated to be 98–99 % due

to the <1 % chance for a mosaic result, defined as some cells

studied showing an abnormality and some cells not showing an

abnormality, and the<1 % chance for maternal cell contami-

nation, defined as studying the mother’s cells instead of the

fetus’ (Ledbetter et al. 1992). CVS may also be used to test for

biochemical abnormalities, single gene conditions, and colla-

gen abnormalities (Pepin et al. 1997; Wapner 2005). CVS

cannot be used to evaluate the risk for ONTDs. The

procedure-related risk of miscarriage is estimated to be approx-

imately 0.5–1%,or1/200–1/100, whether in a singleton or

twin gestation (Canadian Collaborative CVS-Amniocentesis

Clinical T rial Group 1989;CDC,1995; Jackson et al. 1992).

Recent data, however, has shown that the procedure-related

loss rate of CVS may be similar to the rate for mid-trimester

amniocentesis, but this data is only valid in experienced centers

(ACOG Practice Bulletin No. 88, 2007). Of note, there appears

to be an increased risk for limb reduction defects when CVS is

performed prior to 10 weeks gestation (Brambati et al. 1992).

The primary benefit of CVS is that it can be performed at an

earlier gestational age, allowing for earlier decision-making.

Amniocentesis

Amniocentesis is used for the detection of chromosome

aneuploidy in the second and third trimester of pregnancy.

The accuracy for detection of fetal chromosome abnormal-

ities by amniocentesis is estimated to be 99.8–99.9 %

(ACOG Practice Bulletin No. 88, 2007). Amniocentesis

NSGC Practice Guideline 11

may also be used to test for ONTDs, biochemical abnormal-

ities and single gene conditions (CDC, 1995; Winsor et al.

1999). In 1995, the Centers for Disease Control and Preven-

tion released a statement which estimated the procedure-

related risk for miscarriage to be 1/200 or 0.5 % based on

previous studies. Since that time, this number has remained

the universally quoted risk for amniocentesis in the United

States. However, more recent studies actually suggest that

amniocentesis has a much lower risk than the long-accepted

1/200 figure. Data from the First And Second Trimester

Evaluation of Risk for Aneuploidy (FASTER) trial showed

that the miscarriage risk associated with amniocentesis was

approximately 1 in 1600, or less than 0.1 % (Eddleman et al.

2006). In 2007, ACOG released a Practice Bulletin stating

that all women should be offered the option of diagnostic

testing for chromosome aneuploidy, regardless of age, citing

a 1/300 to 1/500 or 0.2–0.3 % risk for miscarriage associat-

ed with amniocente sis based on more recent studies. Many

centers have since adopted this lower, ACOG-e ndorsed

figure (ACOG Practice Bulletin No. 88, 2007). While am-

niocentesis is associated with a lower risk for miscarriage

compared to CVS, one of the primary limitations is the later

gestational age at which the procedure is performed. Waiting

until the third trimester to per form amniocentesis is not

associated with a decreased procedure-related risk; however,

the risk shifts from miscarriage in the second trimester to

preterm delivery in the third trim ester once viability is

reached. Amniocentesis and CVS may be performed in twin

and higher order gestation pregnancies, although the risk for

miscarriage may be increased compared to that associated

with singleton procedures (ACOG Practice Bulletin No. 88).

Early Amniocentesis

Early amniocentesis is available in some centers but is not

typically recommended. It may be performed prior to 15 weeks

of gestation, a nd is associated with an increased risk for

pregnancy loss (2 %), clubfoot (1.3–1.7 %) and fluid leakage

(3.5–4.4 %) compared to routine amniocentesis (<0.1 % for

clubfoot and 1.7 % for leakage), (ACOG Practice Bulletin No.

88, 2007; Canadian Early and Mid-trimester Amniocentesis

Trial, 1998; CDC, 1995; Sundberg et al. 1997).

Screening and Diagnostic Testing Options: Practical

Considerations

Despite numerous potential benefits associated with screening

and testing for chromosome aneuploidy, there are also limita-

tions. Available options may be limited by a number of factors,

including: gestational age of the patient at entry into the health-

care system, state regulations impacting available options, in-

surance coverage and out-of-pocket costs to the patient,

laboratory contracts, availability of laboratory draw sites,

access to certified NT providers, and access to physicians

who perform CVS or amniocentesis. One of the main barriers

faced in screening is insurance coverage, as each private and

public insurance plan has specific requirements for coverage of

screening and diagnostic testing for chromosome aneuploidy.

Additionally, there is often a time lapse between the publication

of guidelines recommending the incorporation of a new test and

routine coverage of that test by insurance companies. These

issues should be considered when assessing a patient’s access to

various prenatal screening and diagnostic testing options.

In addition to external factors that impact testing options,

patient-specific factors also impact decision-making. For

example, patients typically do not view prenatal diagnostic

testing as a routine procedure (Hunt et al. 2005), and may

have conflicting feelings about the possibility of having a

child with a chromosome aneuploidy versus the possibility

of losing a chromosomally normal pregnancy as the result of

a diagnostic procedure (Kupperman et al. 2000). It is up to

the healthcare provider to explore these feelings and discuss

factors that influence decision-making, such as family, so-

cial, and personal history, maternal age, and possible out-

comes in order t o obtain informed consent (Centers for

Disease Control and Prevention 1995). Current practice is

moving away from a maternal age-based risk stratification

for offering diagnostic testing and moving toward custom-

ized risk assessment through the use of the various screen-

ing methods discussed above (ACOG Practice Bulletin

No.88, 2007; Eddleman et al. 2006).

The purpose of the following recommendations is to

assist physicians and allied health professionals in identify-

ing appropriate screening and diagnostic testing options for

chromosome aneuploidy for their patients. Patients may

have differing levels of in ter est in the available options

based on how they feel they might utilize the information.

Recommendations

The information presented above, along with the data sum-

marized in Fig. 1, and Tables 1, 2, 3 and 4 supports the

following recommendations:

Recommendations for all patients

& Providers should offer the options of maternal serum

screening (MSS) and diagnostic testing for chromosome

aneuploidy to every patient.

– Providers should engage in a discussion with their

patients about the benefits, limitations, and risks

of MSS and diagnostic testing so that patients may

make informed and autonomous decisions.

– If the provider feels a patient would benefit from

additional discussion prior to making a decision, a

12 Wilson et al.

referral to a genetic counselor or other qualified

provider may be appropriate.

– Documentation of the patient’s decision to elect

or to decline screeningandtestingshouldbe

made in the patient’s medical record.

– Providers should be aware of factors that may

impact the options available to their patients, such

as the patient’s gestational age, insurance cover-

age and access to services and providers.

& An ultrasound to assess the fetal anatomy is suggested at

approximately 18w0d-20w0d gestation for all patients

regardless of whethe r or not they choose to have screen-

ing or diagnostic testing.

Recommendations for low risk patients less than 14 weeks

of gestation:

& For patients who may consider CVS or amniocentesis,

stepwise sequential screening or combined first trimes-

ter screening should be considered because:

– Both are tailored to fit t he needs of patients

who desire early detection of chromosome

aneuploidy but wish to employ a screening meth-

od prior to making a decision about diagnostic

testing.

– Both allow for the option of CVS in higher risk

pregnancies while deferring testing of lower risk

pregnancies to the second trimester without caus-

ing increased anxiety.

– Of the screening options that provide risk infor-

mat ion in the first trimester, stepwise sequential

screening has the highest detection rates for Down

syndrome and trisomy 18.

& If CVS is not an option, integrated screening may be

considered in order to maximize detection rates.

& If a patient completes combined first trimester screening,

a separate second trimester MSS for chromosom e aneu-

ploidy is NOT indicated. Screening for chromosome

aneuploidy in the second trimester in patients who

present prior to 14 weeks should ONLY be performed

as a part of integrated, serum integrated, stepwise

sequential, or contingency screening. Independent

screening in first and second trimesters increases the

false positive rate of screening.

& Patients who have an increased NT (≥ 95th% or≥

3.0mm) should be offered diagnostic testing by ei-

ther CVS or amniocente sis. A referral for a fetal

echocardiogram should also be considered if the

NT ≥3.5mm.

& Early amniocentesis (pri or to 15 weeks of gestation) is

not recommended due to the increased risks for preg-

nancy loss, clubfoot, and fluid leakage. CVS should be

offered as the diagnostic testing option for chromosome

aneuploidy in the first trimester.

Recommendations for low risk patients after 14 weeks of

gestation:

& Patients who desire MSS but did not have MSS in the

first trimester should be offered a quad or penta screen

rather than a triple screen due to the increased detection

rates.

& Amniocentesis should be offered as the diagnostic test-

ing option for chromosome aneuploidy for patients after

15 weeks of gestation.

Recommendations for patients at increased risk for

chromosome aneuploidy

& Patients who desire screening information may be of-

fered NIPT due to the high detection rates and low false

positive rates. NIPT should only be offered in the con-

text of informed consent, edu cation, and counseling by a

qualified provider, such as a genetic counselor. Standard

confirmatory diagnostic testing should be offered as

follow-up to positive NIPT results. High risk patients

who decline NIPT but remain interested in screening

should be made aware of alternate screening options as

appropriate based on gestational age and screening

availability.

& If the patient presents prior to 14 weeks gestation, CVS

and amniocentesis should both be offered as diagnostic

testing options for chromosome aneuploidy.

& If the patient presents after 14 weeks gestation, amnio-

centesis should be offered as the diagnostic testing op-

tion for chromosome aneuploidy.

Summary

This practice guideline provides a summary of screening

and diagnostic testing options for chromosome aneuploi-

dy and gives realistic recommendations on how to em-

ploy these options. A decision tree and com parison

tables are presented for providers to select the screening

or diagnostic test which best suits their patient’s needs.

Specific recommendations are given for certain clinical

circumstances. However, these options are dependent on

logistical factors such as t iming of entry into healthcare,

insurance coverage, overall cost, and screening/testing

availability. Patients will make decisions regarding these

options based not only on the facts and the data, but

also based on personal feelings, past experiences, and

current perceptions. A referral to a genetic counselor or

other qualified provider may be appropriate if a patient

may benefit from additional discussion prior to making

NSGC Practice Guideline 13