Prenatal diagnosis

Screening versus diagnosis

• Screening (ultrasound markers, maternal serum, circulating fetal DNA) gives a probability. A high-risk result must be confirmed by a diagnostic test before any irreversible decision.
• Diagnostic tests (CVS, amniocentesis, fetal blood sampling) obtain fetal or placental cells for karyotype, rapid aneuploidy testing or DNA analysis.
• Screening is offered to every pregnant woman whatever her age. Maternal age (35 and over was the old cut-off) is only one input into the calculated risk. Counselling is non-directive: the woman decides whether to be tested and what to do with the result.
• Serum results are reported as multiples of the median (MoM) for that gestation, then corrected for maternal weight (a heavier woman has a larger plasma volume, diluting the markers), ethnicity, diabetes, smoking and IVF.

Timeline of tests

First-trimester combined test

• Maternal age + nuchal translucency (NT) + free beta-hCG + PAPP-A. Detects about 85–90% of trisomy 21 at a 3–5% screen-positive rate. NT alone is the best single ultrasound marker.
• Trisomy 21: hCG high, PAPP-A low, NT increased. Trisomies 18 and 13: both low.
• NT of 3.5 mm or more is raised. Even with a normal karyotype the fetus remains at risk of a heart defect (fetal echocardiography is arranged), other genetic syndromes and fetal loss, because the fluid reflects disturbed cardiac and lymphatic development.

Second-trimester serum tests (15 to 20 weeks)

• The triple test omits inhibin A; the quadruple test includes it.
• AFP is made by the fetal yolk sac and liver. It rises with gestation, so the commonest cause of a raised maternal AFP is wrong dates.
• Other causes of high AFP: open neural tube defect, abdominal wall defects, multiple pregnancy, fetal death, feto-maternal haemorrhage, congenital nephrotic syndrome.
• Low AFP: trisomies 21 and 18, overestimated gestation, maternal obesity.

Circulating fetal DNA (non-invasive prenatal testing, NIPT)

• DNA fragments in maternal plasma come from apoptotic placental trophoblast, detectable from about 10 weeks and cleared within hours of delivery.
• Detects about 99% of trisomy 21 with very few false positives, but it is still screening: placental DNA may differ from the fetus (confined placental mosaicism), and a vanishing twin, maternal mosaicism or maternal cancer can give false results.
• Needs a fetal fraction of about 4%. The fraction is lower in maternal obesity and early gestation; a failed test is not a normal test.
• Cell-free DNA also determines fetal RhD status, so anti-D can be targeted.

Invasive diagnostic tests

• The extra miscarriage risk of CVS and of amniocentesis in experienced hands is about 0.5% or less; older texts quote 1–2%.
• Before 14 weeks, transcervical or transabdominal CVS is safer than early amniocentesis, which is why early amniocentesis was abandoned.
• Give anti-D to a rhesus-negative woman after any invasive test.

Processing the sample

• CVS direct preparation: reads spontaneously dividing cytotrophoblast (the inner layer of the villus) within 1–2 days. Long-term culture grows the mesenchymal core, which is closer to the fetus. The syncytiotrophoblast has no dividing nuclei and cannot be karyotyped. Discordance between the two suggests placental mosaicism.
• QF-PCR on uncultured cells counts chromosomes 13, 18, 21, X and Y within about 48 hours, but misses balanced rearrangements and most small deletions.
• A full karyotype needs cell culture (about 2 weeks); chromosomal microarray detects small deletions and duplications and is used when the scan shows a structural anomaly.

Single-gene disorders: thalassaemia in Pakistan

• When both partners are carriers (often a cousin marriage), each pregnancy has a 1 in 4 risk of thalassaemia major.
• The method of choice is CVS at 11–13 weeks with PCR for the known parental mutations. A woman seen at 8 weeks waits until 11 weeks; earlier CVS risks limb defects.
• Karyotype is normal in thalassaemia, and globin chain synthesis studies on fetal blood are only possible later. Identify the parents' mutations first, ideally before pregnancy.
• Pre-implantation genetic testing with IVF tests an embryo cell before transfer and avoids termination.

Neural tube defects and fetal anaemia

• Open neural tube defect on amniocentesis: raised amniotic AFP and an acetylcholinesterase band, which is more specific because fetal blood contamination also raises AFP. A detailed scan (lemon and banana signs) is now usually enough.
• Rhesus disease: a middle cerebral artery peak systolic velocity above 1.5 MoM indicates fetal anaemia (low blood viscosity and high cardiac output speed up flow). This has replaced serial amniocentesis.

Remember Earliest invasive diagnosis is CVS at 11 weeks; amniocentesis waits until 15 weeks.

Exam trap Early CVS risks limb reduction defects; early amniocentesis risks talipes. Do not swap them.

Exam trap A raised maternal AFP is most often wrong dates: confirm gestation by ultrasound before suspecting a neural tube defect.

How FCPS asks about this

• When is CVS done: from 11 weeks (11–13 weeks); amniocentesis from 15 weeks.
• When is nuchal translucency measured: 11 to 13+6 weeks.
• Prenatal diagnosis of beta-thalassaemia in carrier parents: CVS with PCR for the parental mutations.
• Raised NT, normal karyotype: still at risk of a cardiac defect; arrange fetal echocardiography.
• Rapid 48-hour result for 13, 18, 21: QF-PCR on uncultured cells.
• Dividing cells in a CVS direct preparation: cytotrophoblast.
• Quadruple test pattern in Down syndrome: AFP and oestriol low, hCG and inhibin A high.