Genetic amniocentesis is an invasive diagnostic technique.

Common indications are listed in the box below, with prenatal diagnosis being the most common reason.

Chromosomal microarray (CMA) as an adjunct to the standard karyotype is a technique used to detect chromosomal abnormalities (microdeletions and duplications) that are not detected by traditional karyotyping.

Amniocentesis is indicated to obtain fetal material for biochemical or DNA studies.

Molecular abnormalities responsible for many disorders are being identified at an increasingly rapid rate and any listing of these is soon outdated.

A list of the more common genetic conditions for which DNA-based prenatal diagnosis is available is given in Table 3.4.1. Many of these conditions are rare, and their diagnosis is complex, so that consultation with a genetics unit is encouraged before performing an invasive test.

Screening for open spina bifida (OSB) in the second trimester used to be based on the quantification of maternal serum alpha-fetoprotein (MSAFP) concentration in amniotic fluid obtained by amniocentesis; however, it has now been supplanted by ultrasound in many centers (2) because of the advances in ultrasound image quality. Diagnosis has been greatly enhanced by the recognition of associated abnormalities of the skull and brain, including ventriculomegaly, microcephaly, concave deformity of the frontal bones (lemon sign), and obliteration of the cisterna magna with an abnormal anterior curvature of the cerebellar hemispheres (banana sign) (3).

If ultrasound examination demonstrates a normal fetal spine, cranium, and cerebellum, the chance of an undetected open spinal abnormality is low. Amniocentesis can be reserved for patients with suspicious ultrasound findings or large MSAFP elevations despite a normal ultrasound scan, or when there is an inability to adequately visualize fetal anatomy.

Important in women who missed their fetal anatomic survey and in those without the availability of ultrasonographic services, the MSAFP may still play a role in the identification of patients at increased risk for open neural tube defects.

Genetic amniocentesis is commonly performed in the mid-trimester between 15 and 18 weeks’ gestation. At this gestational age, the amount of fluid is adequate (˜150 mL), the ratio of viable to nonviable cells is the greatest, and in most cases, the amnion has fused with the chorion allowing for successful cavity entry, thereby reducing the risk for premature rupture of membranes.

Before the procedure, an ultrasound scan is performed to determine the number of fetuses, to confirm gestational age, to assure fetal viability, to document fetal anatomy, and to localize the placental location and umbilical cord insertion into the placenta.

The maternal abdomen is cleansed with an antiseptic solution (povidone or chlorhexidine). Antibiotic prophylaxis before amniocentesis is not recommended based on a retrospective case-control study (4).

Usually, a 20- or 22-gauge spinal needle is chosen that comes in different lengths. It is important to choose a needle long enough to reach the target pocket accounting for placental location, possible uterine contraction with needle insertion, and maternal abdominal wall thickness. Local anesthesia is generally not necessary as it is uncomfortable and does not abate possible uterine discomfort.

Under direct sonographic guidance, a 20- to 22-gauge spinal needle is introduced into a pocket of amniotic fluid free of fetal parts and umbilical cord (Tech Figure 3.4.1). The pocket should be long enough to allow the advancement of the needle tip into the amniotic fluid cavity. The first 2 cc of aspirated amniotic fluid are discarded to prevent maternal cell contamination and then 20 to 30 cc of amniotic fluid are withdrawn into a sterile syringe and sent for testing. Fetal heart rate should be documented pre- and postprocedure.

Operators are cautioned to identify surrounding maternal bowel and to avoid needle insertion through the bowel and the risks associated with this complication, such as contamination of the amniotic cavity.

Transplacental passage should be avoided when possible, but if unavoidable, the thinnest portion should be traversed. In these instances, color Doppler is instrumental in avoiding maternal or fetal vessels at the sampling site. The area close to the placental cord insertion should be avoided because it contains the largest vessels.

Although blind insertion of the sampling needle was the standard in past years, current expertise has progressed to the point that amniocentesis should only be performed using continuous sonographic guidance. Guidance should be maintained throughout the procedure to avoid inadvertent puncture of the fetus and to identify uterine wall contractions that occasionally will retract the needle tip back into the myometrium (5).

If the initial attempt to obtain fluid is unsuccessful, a second attempt in another location can be performed after reevaluation of the fetal and placental positions. Amniotic membrane tenting and the development of needle-induced uterine wall contractions are the most frequent causes of initial failure.

If the amnion is not fused to the chorion at the initial evaluation for amniocentesis, the procedure should be postponed for a few days to 1 week to improve the chance of obtaining fluid on the first attempt.

Although studies have demonstrated that the fetal loss rate increases with the number of insertions, it does not increase with the number of separate procedures. In experienced centers, return visits are rarely required.

Rhesus-negative women (without alloimmunization to D) should receive prophylactic administration of Rh(D) immune globulin following the invasive procedure.

Amniocentesis in multifetal pregnancy may employ a multineedle technique or a single-needle technique. The multineedle technique involves puncture of the first sac, withdrawal of amniotic fluid, injection of a dilute dye (Indigo carmine) before needle withdrawal from the sac, and then a new needle insertion to puncture the second sac (6,7). If the fluid aspirated after the second puncture is clear, this is confirmation that the first sac was not resampled. If blue-tinged fluid is retrieved, the needle should be removed and another attempt at sampling the second sac should be made.

Methylene blue is associated with skin staining, small-bowel atresia in some studies, and methemoglobinemia; its use is now contraindicated (8). Use of indigo carmine has been reviewed in large series by both Cragan and colleagues (9) and Pruggmayer and colleagues (10), and no increased risk for small-bowel atresia, nor any other congenital anomaly, has been found. However, because of the theoretical risk of an intra-amniotic dye, instillation-free techniques have evolved (5).

Some operators prefer a single-insertion technique. The site of needle insertion is determined by the position of the dividing membrane, and the proximal sac is sampled first (5,7).

After entry into the first sac and aspiration of amniotic fluid, the needle is advanced through the dividing membrane into the second sac. To avoid contamination of the second sample with fluid from the first, the first 2 cc of fluid from the second sample is discarded.

Complications of amniocentesis are rare. A common occurrence following amniocentesis is cramping lasting for 1 to 2 hours. Lower abdominal discomfort may occur for up to 48 hours after the procedure but rarely is it severe. Serious maternal complications such as septic shock are exceedingly rare following amniocentesis.

Amnionitis occurs in <0.1% of cases (1 in 1,000) (1,11) and can occur from contamination of the amniotic fluid with skin flora or from inadvertent puncture of the maternal bowel. It may also follow procedure-induced amnion rupture.

Postamniocentesis chorioamnionitis can have an insidious onset and frequently presents with flulike symptoms with few early localizing signs. This can evolve into a serious maternal systemic infection unless early aggressive treatment is undertaken. Therefore, a high index of suspicion is necessary.

The development of rhesus alloimmunization occurs in ˜1% of Rh-negative women undergoing amniocentesis (12) but can be avoided by prophylactic administration of anti-D immunoglobulin following the procedure.

Amniotic fluid leakage or vaginal bleeding is noted by 2% to 3% of women after amniocentesis and is usually self-limited. Unlike spontaneous second-trimester amnion rupture, fluid leakage following amniocentesis usually resolves after a few days of modified activities and pelvic rest. Occasionally, leakage of amniotic fluid will persist throughout pregnancy (13), but if the amniotic fluid volume remains adequate, a good outcome may be anticipated.

The only prospective, randomized, controlled trial evaluating the safety of second-trimester amniocentesis is a Danish study, which reported on 4,606 low-risk, healthy women, 25 to 34 years old, who were randomly allocated to either amniocentesis or ultrasound examination (14). The total fetal loss rate was 1.7% in the amniocentesis group and 0.7% in the controls (p < 0.01). The observed difference of 1% gave a relative risk of 2.3%.

Miscarriage after amniocentesis was studied in a meta-analysis that estimated a procedure-related loss rate of 0.11% (1 in 900) (16).

The ACOG Practice Bulletin reports a procedure-related loss rate after traditional genetic amniocentesis to be 0.1% to 0.3% when performed by experienced providers (1).

Tabor et al reported the results of a national registry-based cohort study, including all singleton pregnant women who had an amniocentesis (n = 32,852) in Denmark between 1996 and 2006 (17). The authors reported that the number of procedures performed by a department had a significant effect on the risk of miscarriage. In departments performing fewer than 500 amniocenteses, the odds ratio for fetal loss was 2.2 (95% CI, 1.6-3.1) when compared to departments performing >1,500 procedures during the 11-year period.

In early experience with amniocentesis, needle puncture of the fetus was reported in 0.1% to 3.0% of cases (18). Continuous use of ultrasound to guide the needle minimizes needle puncture of the fetus, and in experienced centers, this is an exceedingly rare complication.

Except for hemolytic disease caused by alloimmunization, no long-term adverse effects have been demonstrated in children undergoing amniocentesis.

Women interested in genetic amniocentesis known to be infected with hepatitis B, hepatitis C, or human immunodeficiency virus need additional counseling about the risk of vertical transmission. With chronic hepatitis B, genetic amniocentesis does not appear to increase the risk of neonatal infection with hepatitis B. These neonates generally receive the hepatitis B vaccine and immune prophylaxis (HBIG) after delivery. Women with HIV who are not on highly active antiretroviral therapy (HAART) and women with high viremic loads of hepatitis C should be counseled on noninvasive screening methods as data on the risk of vertical transmission, especially if the procedure is transplacental, are limited (1).

Compared to singletons, the loss rates for twins before 28 weeks are somewhat higher. Most series report postprocedure loss rates between 2% and 5% (Table 3.4.2).

Although the higher loss rates could demonstrate an increased danger of amniocentesis in twins as compared to singletons, it more likely is a manifestation of the inherent risk of twin gestations.

Patients with twins must also be counseled about the risk of finding a karyotypically abnormal child, which, because of the presence of two fetuses, is approximately twice that following a singleton procedure (19).

Families need to consider the possibility of a test showing that one of the twins is normal and the other has an abnormality. Selective termination of the affected fetus is an option; however, it is associated with a postprocedure loss rate of 5% to 10% (20). However, this approach is also associated with an increased risk of preterm birth, especially when performed after 20 weeks gestation or if the lower fetus is terminated (21).

The major drawbacks of conventional second-trimester genetic amniocentesis are the delayed availability of the karyotype and the increased medical risks of a dilatation and evacuation procedure late in the pregnancy.

Delaying the procedure until after fetal movement is felt imposes a severe emotional burden on the patient. Because of these concerns, CVS is now offered in most referral centers.

CVS, which samples the placental villi rather than the amniotic fluid, is one of the most successful approaches to date for moving prenatal diagnosis into the first trimester of pregnancy.

The recent introduction of fetal DNA testing in the maternal blood may influence the number of CVS procedures performed at referring centers; however, fetal DNA testing is not considered diagnostic at present.

The conventional approach is to offer CVS between 11 and 14 weeks of gestation. Most spontaneous pregnancy losses would have occurred by 11 weeks; thus, by postponing CVS at or beyond 11 weeks, fewer CVS procedures may be needed.