Fetal Medicine

First-trimester combined screening test

In the United Kingdom, screening for chromosomal anomalies is offered to all eligible pregnant women to assess the chance of the baby being born with Down syndrome (trisomy 21 or T21), or Edwards syndrome (T18) or Patau syndrome (T13).

The test of choice is first-trimester combined screening. Women can choose:

• 1

  Not to have screening

• 2

  To have screening for T21 and T18/T13

• 3

  To have screening for T21 only

• 4

  To have screening for T18/T13 only

The combined screening test is performed between 11 ⁺² weeks to 14 ⁺¹ weeks of gestation, which corresponds to a crown rump length (CRL) of 45.0 mm to 84.0 mm ( Fig. 25.1A ). To calculate the chance of the pregnancy being affected by T21 or T18/T13, the combined test incorporates:

• Maternal age

• The measurement of the thickness of nuchal fluid behind the fetal neck (nuchal translucency [NT]; Fig. 25.1B )

• Maternal biochemistry, specifically free β‐human chorionic gonadotrophin (β‐hCG) and pregnancyassociated plasma protein A (PAPP-A)

• The gestational age calculated from the CRL measurement.

This combination of tests has a detection rate of 85% for Down syndrome when using a threshold risk of 1 in 150 for the definition of ‘high chance’. This is associated with a 3% false-positive rate. In dichorionic twin pregnancies, each twin is given an ‘individualised’ chance. In monochorionic twins, the chance is calculated as an average of that allocated to both twins. As previously discussed, these screening tests can identify only those fetuses at increased risk of problems. For absolute certainty regarding whether a fetus is affected, an invasive diagnostic test will be required (see the amniocentesis and chorionic villous sampling [CVS] section later in the chapter).

Second-trimester quadruple test

For those in whom it is not possible to obtain a first-trimester screening test (such as when an NT measurement cannot be obtained in the first trimester due to poor fetal position or a pregnancy that is already beyond 14 weeks’ gestation), a serological test for Down syndrome may be offered (but not for T18 or T13). The quadruple test incorporates maternal age and four biochemical markers measured between 14 ⁺² weeks until 20 ⁺⁰ weeks: alpha-fetoprotein (AFP), free β‐hCG, estriol, and inhibin-A. The detection rate is 80%, lower than for the combined screening test (85%), with an associated 4% false-positive rate. Down syndrome is associated with low levels of AFP and unconjugated estriol and high levels of free β‐hCG and inhibin A. A raised AFP should also prompt an ultrasound scan looking for a neural tube defect or gastroschisis (see later discussion).

Non-invasive prenatal screening test (NIPT)

NIPT is the analysis of cell-free fetal deoxyribonucleic acid (cffDNA) in maternal serum to screen for trisomies 21, 18, and 13. These are fragments of DNA that are released from the placenta into the maternal circulation. NIPT may be offered from 10 weeks of gestation, as by this stage, cffDNA makes up an average of 10% of cell-free DNA in the mother’s blood and is detectable at serum testing.

NIPT is a far more sensitive and specific screening test than either the combined or quadruple test. However, it is more expensive and, therefore, is currently recommended by the NHS as an option for women who have been identified with a ‘high chance’ (>1 in 150) by either first-trimester combined or quadruple tests but who wish to avoid invasive diagnostic procedures. The detection rate for T21 is 99.7%, with an associated 0.01% false-positive rate. The detection rates for T18 and T13 are 97.4% and 93.8%, respectively, associated with a 0.13% false-positive rate. It is important to emphasise to women that NIPT is still a screening test, and those that are deemed to have a ‘high chance’ of T21, T13, or T18 are advised to undergo a diagnostic invasive test as detailed later.

There are several private providers offering NIPT screening for conditions above trisomies 21, 18, and 13. Some providers offer fetal sexing; screening for sex chromosome aneuploidies, such as Turner (45 X) and Klinefelter (47 XXY) syndromes; or microdeletions and duplications, such as DiGeorge syndrome (22q11 deletion). These indications are ethically controversial, lack scientific validation, and not supported by professional societies.

As cffDNA is produced by the placenta rather than the fetus, the accuracy of NIPT is influenced by a variety of factors. NIPT accuracy is reduced in twin pregnancies (NIPT cannot be offered to higher-order multiples), obesity, and at less than 10 weeks’ gestation (due to insufficient fetal DNA in the maternal circulation). False-positive results can also be caused by confined placental mosaicism or a ‘vanishing twin’ (a twin pregnancy in which demise of one twin results in a singleton pregnancy) and maternal malignancy.

Chorionic villus sampling (CVS)

CVS is typically performed between 11 and 14 weeks’ gestation and involves passing a needle transabdominally, or occasionally transvaginally, under ultrasound guidance by an appropriately trained fetal medicine specialist to take a sample of placental tissue (villi). The risk of miscarriage is commonly reported to be 0.5%. At this early gestation, it is difficult to determine whether the miscarriage would have happened anyway due to an underlying genetic defect in the fetus or whether it is the result of the CVS.

Amniocentesis

Diagnostic amniocentesis may be performed from 15 weeks’ gestation. It involves passing a thin needle transabdominally into the amniotic cavity, under continuous ultrasound guidance, to extract 10 to 18 mL of amniotic fluid. The risk of miscarriage is reported to be the same as CVS, at around 0.5%.

Genetic testing

Traditionally, chorionic villi or amniocyte (fetal fibroblasts) culture was used to obtain karyotype results for both CVS and amniocentesis. This has been superseded by the following newer tests. Quantitative fluorescence polymerase chain reaction (QF-PCR) provides a more rapid assessment of the more common aneuploidies. Fluorescently labelled markers are used to amplify specific regions of DNA to quantify the amount of DNA present in those regions. Results are available within 48 hours and are limited to the detection of T21, T13, T18, and 45X (Turner syndrome). Chromosome microarray analysis (CMA) has mostly replaced traditionally karyotyping. CMA provides a more detailed assessment of alterations in the genome. It can detect small gains and losses of genetic material, known as ‘copy number variants’ (CNVs). If fetal anomalies are present, microarray provides an additional diagnostic yield of 6% when the karyotype is reported as normal. Detected CNVs may be known to be pathogenic or, in up to 2% of cases, may be variants of unknown clinical significance (VOUS). In view of uncertainty, VOUS may lead to unnecessary parental stress and anxiety.

Whole-genome and exome sequencing for prenatal diagnosis are being explored largely in research settings and in highly specialised clinical genetics units. Whole-genome sequencing (WGS) involves sequencing or ‘reading’ the entire fetal genome. Exomes refer to the 1% to 3% of the genome that contains the vast majority of active genes. Whole exome sequencing (WES) therefore provides a more efficient yet still comprehensive genome assessment.

The NHS is the first national health care system to offer WES, providing an additional diagnostic yield of 8% to 10% when the microarray is reported as normal. At present, the utility of WGS and WES is limited by significant cost and the limited ability to correlate the significance of observed genomic variations with the phenotype being evaluated. As genome reference libraries expand, so will the capabilities of WGS and WES for prenatal diagnosis.

Non-invasive prenatal diagnosis (NIPD)

NIPD is based on analysis of cffDNA circulating in the maternal plasma to identify specific monogenic disorders. Monogenic disorders affect around 1% of us; there are a few common scenarios when single-gene testing is used for prenatal diagnosis. This is undertaken when there is a known familial mutation for which the fetus is at risk or when prenatal ultrasound examination identifies features that are highly characteristic for a well-defined specific disorder.

NIPD is now offered for a small number of single-gene disorders, including achondroplasia, thanatophoric dysplasia, Apert syndrome, congenital adrenal hyperplasia, and cystic fibrosis. It can also be used for fetal sex determination when the pregnancy is at risk of a sex-linked disorder. NIPD offers definitive diagnosis and does not require an invasive test for confirmation of diagnosis, eliminating the risks associated with invasive procedures.

NIPD is most commonly employed to diagnose fetal rhesus D-antigen status for rhesus D-negative mothers. A fetus predicted to be rhesus D-negative by NIPD precludes the need for antenatal anti-D prophylaxis in the mother.

Down syndrome (trisomy 21)

The overall incidence is approximately 1 in 1000 live births; however, this is known to increase with advancing maternal age:

• 20 years, 1 in 1500

• 30 years, 1 in  800

• 35 years, 1 in  270

• 40 years, 1 in 100

• 45 years, 1 in  50

All children born with Down syndrome (or Down’s syndrome) have some degree of neurodevelopmental delay; however, this is extremely variable. Likewise, the physical characteristics commonly associated with the condition – including hypotonia, short stature, facial features with flat nasal bridge and protruding tongue, upward slanting eyes, broad hands and single palmar crease – manifest to differing degrees. Around 50% of those born with Down syndrome will also have a congenital heart defect, most commonly a septal defect. Down syndrome is also associated with gut mobility problems, hypothyroidism, and an increased risk of early-onset dementia. Life expectancy for those affected with Down syndrome has increased dramatically in the last 50 years, from 25 in the 1980s to 60 today. Of all cases, 95% are due to chromosomal nondisjunction, with 4% due to translocation, and the remaining 1% to mosaicism.

Edwards syndrome (trisomy 18)

The incidence is around 1 in 3500 live births, with the majority resulting from nondisjunction of chromosome 18. Clinical presentation is characterised by early-onset growth restriction, specific craniofacial features, including a small strawberry-shaped cranium, small facial features and low-set ears, and skeletal abnormalities, including overlapping fingers and prominent calcanei (rocker bottom feet). Major systemic abnormalities are common and include congenital heart disease, complex urogenital anomalies, and problems with the gastrointestinal system, such as omphalocele and oesophageal atresia. Approximately 68% die in utero; for those who survive to birth, the outcome is poor. Median survival is 2 weeks; up to 13.5% live to 1 year and 12.3% to 5 years.

Patau syndrome (trisomy 13)

The incidence is low, ranging from 1 in 6500 to 1 in 29,000 live births. It is associated with multiple severe congenital abnormalities, which result in significant physical and mental impairment. Approximately 80% of children will have a severe heart defect. Significant brain defects, clefts of the lip or palate, kidney and urogenital anomalies, extra digits, omphalocele, and spina bifida are also common features. The majority of babies affected are stillborn, with survivors rarely expected to live longer than 1 week. Only 11.5% will survive the first year.

Triploidy

This describes the presence of an additional set of chromosomes acquired either from the mother (causing severe fetal growth restriction and fetal abnormality) or father (associated with a partial mole; see Chapter 15) during fertilization, resulting in a total of 69 as opposed to the normal 46. Affected fetuses usually miscarry in early pregnancy and survival to birth is rare. There is no expected survival past the immediate neonatal period, with the affected fetus generally severely growth restricted and affected by multiple severe abnormalities.

Turner syndrome (45,XO)

This affects around 1 in 2500 live-born girls. In most cases, it is due to the loss of the paternal chromosome, although some individuals have a mosaic pattern. Antenatally, Turner syndrome is associated with cystic hygroma ( Fig. 25.3 ), cardiac defects, and non-immune hydrops, which results in many affected pregnancies miscarrying. If not identified antenatally, the majority of girls affected are diagnosed in infancy or childhood as a result of characteristic physical features. These include short stature, webbed neck, widely spaced nipples, and cubitus valgus. Other associated problems include renal dysgenesis, coarctation of the aorta, and ovarian failure, necessitating long-term hormone replacement therapy (HRT) requirements. Intelligence is largely unaffected, although there may be some impairment of non-verbal skills.

47,XXX

This is the most common female chromosomal abnormality, occurring in 1 in 1000 live births, although higher in pregnancies in women over 40 years. Those affected are phenotypically normal, with normal development of secondary sexual characteristics and fertility. There is often a delay in motor and speech development and an association with genitourinary problems, including premature ovarian insufficiency, requiring HRT.

Klinefelter syndrome (47,XXY)

This affects 1 in 1000 live births and is a frequent cause of male factor infertility. Those affected tend to be tall males with sparse body hair and gynaecomastia. Typically, the testes remain small; many cases are diagnosed during puberty as a result of this. There is some association with reduced intelligence quotient (IQ), hypothyroidism, cardiovascular disease, and type 2 diabetes.

Jacobs syndrome (47,XYY)

Incidence is around 1 in 1000 live births and is frequently undetected due to a lack of symptoms or fertility concerns. Characteristically, males are tall with acne. Whilst intelligence is in the normal range, there may be an association with behavioural problems, including impulsivity.

Haemoglobinopathies

Thalassaemias and sickle cell disease are recessively inherited genetic conditions of the haemoglobin gene, which have serious health implications and require lifelong treatment. The NHS Sickle Cell and Thalassaemia Screening Programme recommends that all pregnant women be offered a blood test by 10 ⁺⁰ weeks of gestation to determine whether they carry a gene for thalassaemia and to differentiate those at high risk of being a sickle cell carrier. In low-prevalence areas, the family origin questionnaire (FOQ) information is used as an initial screening tool to assess a woman’s eligibility for haemoglobin variant screening. All biological fathers are offered screening if the pregnant woman is a genetic carrier for sickle cell disease or thalassaemia.

When both parents are carriers, in each pregnancy, the risks to the baby are:

• 1 in 4 (25%) chance of being completely unaffected

• 2 in 4 (50%) chance of being a carrier

• 1 in 4 (25%) chance of inheriting the condition

Couples/women at risk of having a baby with sickle cell disease or thalassaemia major are referred to health care professionals who provide counselling and prenatal diagnosis. The diagnostic approach usually depends on the gestational age of the fetus. The fetal sample for prenatal diagnosis is usually obtained by CVS or amniocentesis.

Cystic fibrosis

This is an autosomal-recessive condition, with a UK incidence of around 1 in 2500. It is caused by mutations in the cystic fibrosis transmembrane conductance regulator gene on chromosome 7, which codes for a protein involved in chloride ion channel function. Alterations in this protein can lead to thickened mucous affecting the lungs, resulting in recurrent chest infections, pancreatic insufficiency, and malabsorption. Azoospermia in males is common, with subsequent subfertility. The prognosis in cystic fibrosis has significantly improved with the development of new drug treatments such as the combination of tezacaftor, ivacaftor and elexacaftor. If a couple are known to be carriers, there is a 25% risk of their baby being affected. Thus, it is reasonable to offer NIPD (assuming that both parents are CF carriers and have DNA from an affected or unaffected child) or invasive testing if they felt that they would not continue with an affected pregnancy.

Huntington disease

This is an autosomal-dominant condition with a peak age of onset between 40 and 45 years. It is caused by a CAG trinucleotide expansion and can result in chorea, dementia, and neuropsychiatric disturbance. Generally, those affected deteriorate over time, and life expectancy tends to be about 20 years from the onset of symptoms. Given the lack of treatment, and the fact that the disease tends to manifest in later life, the ethical issues around testing the children of sufferers are complex. At present, testing is offered to those over 18 years who wish to proceed following genetic counselling. There is also the option for invasive prenatal testing, although this becomes even more complex if a member of the couple is at risk but does not wish to know.

DiGeorge syndrome

This is due to a deletion in a small part of chromosome 22 (22q11 deletion). It affects 1 in 350 pregnancies and 1 in 2000 live births. Most are due to de novo mutations. Ultrasound findings include congenital heart abnormalities, kidney defects, and facial features, including a cleft palate. Those affected have a poor immune system, a higher incidence of learning difficulties, and, longer term, 1 in 4 will develop schizophrenia.