Amniocentesis

Definition

Amniocentesis is an optional but reliable procedure offered to pregnant women in order to obtain more information about a developing fetus in the second trimester of pregnancy. It is primarily offered to pregnant women who are at increased risk, based on their age, family history, or other factor, of having a child with a genetic condition. Amniocentesis provides accurate information about fetal chromosomes or the likelihood of certain physical abnormalities. Additional specialized studies may be performed on an as-needed basis. Women who undergo amniocentesis typically do so either to obtain reassurance about fetal well-being or, if the results are abnormal, to plan for the remainder of their prenatal care. The procedure is associated with a slightly increased chance for pregnancy loss.

Purpose

Amniocentesis has been considered a standard of obstetrical care since the 1970s. It is not, however, offered to all pregnant women. The American College of Obstetricians and Gynecologists (ACOG) recommends that amniocentesis be offered to all expectant mothers age thirty-five and older. This age cut-off has been selected because advancing maternal age is associated with an increasing risk of having a baby with a numerical chromosome abnormality. At age 35, this risk is approximately one in 200 births roughly equivalent to the risk of pregnancy loss associated with amniocentesis.

Amniocentesis is performed for the following reasons:

• Detection of structural chromosome abnormalities or an increased number of chromosomes in the fetus when the mother is in a high risk group. This includes a maternal age of 35 or more; a mother who is known to carry an abnormal chromosome such as a 21/14 translocation; family history of chromosome abnormalities; a genetic disease such as sickle cell anemia for which a DNA test is available; or abnormal triple marker screening test for Down syndrome (i.e., a low serum alpha fetoprotein or estriol or a high chronic gonadotropin level).
• Determination of the sex of the fetus when the mother is a carrier of an X-linked genetic disease such as hemophilia.
• Confirmation of an elevated maternal serum alpha fetoprotein result. Alpha fetoprotein is increased in amniotic fluid in pregnancies associated with open neural tube defects such as spina bifida and several other abnormalities that allow the fluid to leak from the fetus into the amniotic fluid.
• To evaluate the severity of hemolytic disease of the newborn (HDN). In this condition, maternal antibodies, usually anti-Rho (D), cross the placenta and facilitate destruction of Rh positive fetal red blood cells.

• Evaluation of fetal lung maturity (FLM). Respiratory distress syndrome results from collapse of the infant's lungs during expiration if the level of pulmonary surfactants is insufficient. This may occur immediately after birth especially in premature deliveries and when the mother is a diabetic. Measurement of surfactants excreted by the fetus into the amniotic fluid is used to determine the maturity of the fetal lungs.

Evaluation studies of HDN and FLM are performed in the second half of pregnancy, whereas tests for alpha fetoprotein (AFP), chromosome karyotyping, and abnormal genes are performed during the first half of pregnancy, typically between weeks 15 and 18.

Precautions

Amniocentesis is associated with a slightly increased chance of pregnancy loss (approximately 0.5%). Each woman should discuss the potential risks and benefits of amniocentesis with a doctor or genetic counselor and make her own decision about whether or not she desires this testing.

Early amniocentesis, or procedures performed before the thirteenth week of pregnancy, has been associated with an increased risk of clubfoot and of procedure-related pregnancy loss. When collecting amniotic fluid universal precautions should be followed for the prevention of transmission of bloodborne pathogens. Bloody fluid or fluid containing meconium may interfere with the tests for amniotic fluid bilirubin performed to evaluate the severity of HDN. These substances may also interfere with the interpretation of some FLM tests such as the L:S ratio (which is used to determine the level of the fetus's lung development). Some tests such as the amniotic fluid bilirubin test and alpha fetoprotein test must be interpreted with regard to gestational age.

Description

According to the National Center for Health Statistics (NCHS), 112,776 amniocentesis procedures were performed in the United States in 1998, the most recent year for which data is available. The annual birth rate that year was approximately 3.9 million infants. Thus, approximately 3% of pregnant women in the United States had this procedure performed. It is likely that this is an underestimate. The NCHS obtains information from birth certificates registered in each state and the District of Columbia. Although almost all deliveries are registered in the United States, records are still submitted with incomplete information. It is also not possible to know how many amniocentesis procedures were performed for genetic testing, as compared to other indications, as this information is not requested.

Normal persons have a total of 46 chromosomes in each body cell, with the exception of sperm or egg cells that should have only 23. As women get older, there is an increased risk of producing an egg cell with an extra chromosome. This leads to an egg cell with 24 chromosomes rather than the normal 23. Pregnancies with an abnormal number of chromosomes are referred to as aneuploid. Aneuploidy results in a conceptus with either too much or too little genetic material. This, in turn, leads to abnormal development. Aneuploidy results from failure of the chromatids to separate either during meiosis or in the germ line cells of the embryo. Common effects of aneuploidy include an increased risk for pregnancy loss or, in living fetuses, for mental retardation and physical abnormalities.

Down syndrome is the most common form of aneuploidy in live born infants, occurring in approximately one in 800 births, regardless of maternal age. The most common form of Down syndrome is 21 trisomy. One of the gametes contains an extra chromosome number 21 resulting in three number 21 chromosomes following fertilization. In women who are 35 years old, the risk of having a child with Down syndrome is higher, roughly one in 385 at delivery. It is important to realize that Down syndrome is not the only chromosome abnormality that may occur. Other numerical abnormalities are possible, yielding genetic conditions that may be either more or less severe than Down syndrome. Thus, a woman is often at risk, based solely on her age, of having a child with any type of chromosome abnormality. At age 35, this total risk is approximately one in 200. By age 40, this risk has increased to one in 65, and, at age 45, this risk is one in 20. These numbers reflect the risk at the time of delivery.

Women younger than 35 years may also have children with chromosomal or other genetic disorders. Therefore, other indications for amniocentesis or other forms of prenatal diagnosis include a family history of or a previous child with a known genetic condition, abnormal prenatal screening results, such as ultrasound or a blood test, or one parent with a previously identified structural chromosome rearrangement. All of the above may make it more likely for a couple to have a child with a genetic condition.

Amniocentesis is the most common invasive prenatal diagnosis technique offered to pregnant women. Its safety and accuracy are well-established, and it is generally considered the standard by which other prenatal diagnosis techniques are measured. To perform the procedure, a doctor inserts a thin needle into the mother's uterus and the amniotic sac. A continuous ultrasound evaluation is typically used so that the doctor can avoid touching either the baby or the umbilical cord with the needle. The amniotic sac is made up of two membranes: the inner amnion and the outer chorion. The amnion and chorion both develop from the fertilized egg. They are initially separate but begin to fuse early in pregnancy. This fusion is usually completed by approximately the fourteenth to fifteenth week of pregnancy.

Amniocentesis is usually performed in the second trimester, usually during weeks fifteen to eighteen (mid-trimester). The amniotic sac holds the fetus suspended within the amniotic fluid, an almost colorless fluid that protects the fetus from harm, helps maintain a consistent temperature, and prevents the fetus, or parts of it, from becoming attached to the amnion. The amniotic fluid is produced and absorbed by the fetus and the mother throughout pregnancy. Fetal cells, primarily derived from the skin, digestive system, and urinary tract, are suspended within the fluid. A smaller number of cells from the amnion and placenta are also present. Finally, the fetus produces a number of different chemical substances that also pass into the amniotic fluid. One of these substances is bilirubin, a breakdown product of hemoglobin. Bilirubin in amniotic fluid is measured to determine the extent of fetal red cell destruction in HDN. Pulmonary surfactants are used in some higher-risk pregnancies to assess fetal lung maturity, and alpha-fetoprotein is used to screen for certain structural birth defects.

It is possible to perform amniocentesis in a twin pregnancy. Amniocentesis in some higher-order pregnancies, such as triplets, has also been reported. In a multiple pregnancy, it is important to ensure that a separate sample of amniotic fluid is obtained from each fetus. To accomplish this, a doctor injects a small amount of harmless blue dye into the amniotic sac of the first baby after a sample has been withdrawn. The dye will temporarily tinge the fluid blue-green. A second needle is inserted into the next amniotic sac with ultrasound guidance. If the fluid withdrawn is pale yellow, a sample from the next fetus has been successfully obtained. Since monoamniotic twins share the same amniotic sac, the second sample will be blue.

Chorionic villus sampling

Mid-trimester amniocentesis has been available for nearly thirty years. Chorionic villus sampling (CVS) has been available in the United States since the 1980s. CVS is usually performed between ten and twelve weeks of pregnancy. It involves the removal of a small sample of the developing placenta, or chorionic villi. It has been an attractive alternative to amniocentesis, particularly for those women who desire both testing and results earlier in their pregnancies. Some of the benefits of earlier testing include reassurance sooner in pregnancy and fewer physical complications following first trimester pregnancy termination, for those couples with abnormal babies who choose this option. CVS is, however, associated with a higher risk of miscarriage than mid-trimester amniocentesis and because it is performed earlier than amniocentesis more fatal chromosome abnormalities are detected that would not come to term. At experienced centers, this risk is approximately 1% (or one in 100).

Early amniocentesis

Early amniocentesis is performed before the thirteenth completed week of pregnancy. It has been considered investigational for many years. The results of the largest early amniocentesis trial, published in 1998, have, however, caused physicians worldwide to reconsider the benefit and risks of this earlier procedure. The Canadian Early and Mid-trimester Amniocentesis Trial (CEMAT) is the largest, multi-center, randomized clinical trial of early amniocentesis conducted to date. The purpose of the trial was to examine and compare the safety and accuracy of early amniocentesis (EA) versus mid-trimester amniocentesis (MTA). In order to accomplish this, 4,374 pregnant women were identified and enrolled in the study. Ultrasound was performed in the first trimester to confirm the gestational age of all pregnancies. Computer randomization was used to evenly divide the women into either the EA or MTA groups. Ultimately, 1,916 women underwent EA and 1,775 women had MTA. Follow-up was obtained on nearly all pregnancies. Two striking conclusions were reached: EA is associated with an increased incidence of clubfoot and an increased risk of procedure-related pregnancy loss. EA was also linked to an increased number of laboratory culture failures (no growth of cells and no results) compared to MTA. The total waiting time for results was slightly longer in the EA group. This is not entirely a surprise. A smaller amount of fluid is obtained when EA is performed. Hence, there are fewer cells, and culture times take longer.

Chromosome tests

Genetic testing is available on amniotic fluid obtained by amniocentesis. The most common test is a complete analysis of the fetal chromosomes called karyotyping. This procedure detects only those chromosome abnormalities that produce either a structurally abnormal chromosome (e.g., a long or short arm deletion) or aneuploidy. It does not detect mutations within genes that are responsible for specific diseases or inborn errors of metabolism (e.g., cystic fibrosis ). However, many DNA based tests are available that will detect disease genes that are caused by point mutations (single base substitutions) within the genes.

After a sample of amniotic fluid is obtained, the cytogenetic laboratory isolates the cells, referred to as amniocytes, out of the fluid. The cells are placed into two or more containers filled with liquid nutrients, establishing different cultures in which the cells will continue to grow. The cells are cultured anywhere between seven to 10 days before the actual analysis begins. Culture time can be reduced to five to six days by growing a monolayer of fetal cells directly onto a coverglass. Culture is performed in order to obtain sufficient metaphase cells and to synchronize the growth of the cells. Cells in metaphase are selected for analysis because at this stage the chromatids have duplicated but not separated and the individual chromosomes are most visible under the microscope.

Once there appears to be an adequate number of cells to study, the cultures are harvested. Harvesting prevents additional cell growth and stops the cells at whatever point they were in the cell cycle. Cells are washed, exposed to hypotonic saline, and fixed with methanol and acetic acid. The cells are spread on a glass slide, heat fixed and treated with trypsin and stained with Giemsa or other chromosome stain. Smears are examined under 400x magnification and the modal chromosome count is recorded. Large clear metaphase cells are selected for photography or image analysis using chromosome-sorting software. Chromosomes are classified by size, centromere position, banding pattern, secondary constriction, and satellite appendages. The karyotype or chromosome map that results identifies translocations, aneuploidy, deletions, and other abnormalities in the appearance of the chromosomes. Typically, chromosome results are available within seven to 14 days after amniocentesis. Results may be delayed by slow-growing cultures. This rarely reflects an abnormal result but does extend the time until final results are available.

Many laboratories now perform a technique called fluorescence in situ hybridization (FISH) into their chromosome studies. This adjunct testing provides limited information about certain chromosomes within one to two days after amniocentesis. It does not replace a complete chromosome study using amniocyte cultures. In fact, FISH results are often reported as preliminary, pending confirmation by cultured results. They can, however, be very useful, particularly when there is already a high level of suspicion of a fetal chromosome abnormality.

FISH is performed using a small sample of uncultured amniotic fluid cells. Special molecular tags consisting of single-stranded DNA that are attached (covalently conjugated) to fluorescent dyes are used. Each tag is bound to a different combination of dyes. When excited by ultraviolet light, the fluorochromes emit visible light of characteristic colors. Tags are available that recognize a specific segment of DNA from chromosomes 13, 18, 21, X, and Y. Abnormalities of these five chromosomes account for nearly 95% of all chromosomal abnormalities. The cells are prepared onto slides as described for karyotyping, denatured to separate the DNA strands, and the probe mixture is added. Each probe hybridizes with the complementary DNA sequence of the target chromosome. When the fetal cells are examined under a fluorescent microscope, a small spot of a specific color will appear on each of these chromosomes. If an extra autosomal chromosome is present, three colored spots appear within the nucleus of the cell instead of two. In addition to determining sex, FISH detects extra sex chromosomes such as 47,XXY which is responsible for Klinefelter syndrome or the absence of one sex chromosome, and 45,X0 which is responsible for Turner syndrome. Other chromosomal abnormalities will not be detected unless a full chromosome evaluation on cultured cells is performed.

A new fluorescent mapping technique called spectral karyotyping (SKY) is available to aid in identifying chromosomes. A DNA probe specific for each chromosome is tagged with between one to five fluorescent dyes. Hybridization between the tag and DNA of the specific chromosome produces a unique color for each one. Using fluorescent microscopy, and a color camera, the karyotype showing each of the 24 different colored chromosomes is constructed. Chromosomes in which translocations occur are bicolored. The color coding facilitates the identification of the regions of each of the chromosomes involved.

DNA tests are offered to those couples who, based on their family history or other information, are at increased risk of having a child with a single gene, or Mendelian, disorder. Testing for disease genes is primarily accomplished using the polymerase chain reaction (PCR). This technique permits a small segment of DNA within a gene to be amplified exponentially. Within two hours, the target DNA sequence can be copied over one million times, yielding sufficient DNA to identify with a radiolabeled or enzyme-conjugated probe. An example of this technique has been applied extensively to the prenatal diagnosis of cystic fibrosis. There are over 200 different point mutations identified in the CF gene, a segment of 27 coding regions called exons. Various probes may be used to identify most of these. For example, approximately 85% of all CF genes can be identified in the North American white population. DNA tests are available for sickle cell disease, muscular dystrophy, retinoblastoma, hyperlipoproteinemia, Huntington's chorea, Tay-Sachs disease, congenital adrenal hyperplasia, hemophilia, and many other conditions.

Biochemical testing of amniotic fluid is performed using the same methods as are applied to other body fluids. For example, fluorescence polarization and thin layer chromatography are techniques used to measure pulmonary surfactants. Bilirubin in amniotic fluid is analyzed by direct absorbance measurement using a scanning or diode array spectrophotometer. Bilirubin absorbs light maximally at 450 nm, and if present in the amniotic fluid, it will produce an absorbance peak at this wavelength. Hemoglobin contributes to the absorbance peak by also absorbing light at this wavelength. The interference from hemoglobin can be corrected by measuring its absorbance peak at 410 nm. The contribution of hemoglobin to the 450 nm peak is equal to 5% of its absorbance at 410 nm. Therefore, the 5% of the absorbance at 410 nm is subtracted from the absorbance at 450 nm to determine the corrected absorbance which is proportional to bilirubin concentration.

Alpha fetoprotein in amniotic fluid is measured by a sandwich type enzyme immunoassay, the same method used for maternal blood. AFP is a protein made by the fetal liver. It passes out of the fetus and enters both the amniotic fluid and the mother's blood. Screening for open neural tube defects, abnormal openings in the fetal head or spinal cord, or ventral wall defects, openings along the abdominal wall, can be performed by measuring AFP during the fifteenth to twentieth weeks of pregnancy. An unusually high level of serum AFP does not necessarily indicate a problem with fetal development, but is cause for some concern. A high AFP level in amniotic fluid will detect up to 98% of all openings on the fetal body that are not covered by skin. Further studies may be suggested if the AFP is high. Most initial AFP results are available within two to three days after amniocentesis.

Preparation

The usual precautions concerning sterility of the injection site and sterility of equipment and the hands of those who will come into contact with a woman must be taken during amniocentesis.

Counseling must be offered to a woman or couple to ensure that they fully understand the results of an amniocentesis test. Counseling should be offered prior to the procedure so that a woman or couple can understand the procedure and slight risk of miscarriage that is associated with amniocentesis.

Aftercare

Physical care after an amniocentesis procedure involves the site of needle insertion. It should be kept clean and dry. Covering the puncture site with a sterile bandage is usually sufficient.

Mental care after an amniocentesis may be more important than physical care. Many women or couples require emotional support and reassurance while they are waiting for test results. Those receiving abnormal results often require ongoing support while they weigh options and make decisions that are appropriate for them. Some may require long-term support as they adjust to the outcomes of their decisions.

Complications

Women who have had an amniocentesis often describe it as uncomfortable, involving some mild pressure or pain as the needle is inserted. Fewer women describe it as extremely painful. A local anesthetic may be used to numb the upper layer of the mother's skin prior to testing. This medicine has no effect on the fetus, but may help the mother feel more comfortable during the procedure. An experienced physician can, on average, perform amniocentesis in approximately one to two minutes.

Common complaints after amniocentesis include mild abdominal tenderness at the site of needle insertion or mild cramping. These usually go away within one to two days. More serious complications are significantly less common but include leakage of amniotic fluid, vaginal bleeding, or uterine infection. These complications are estimated to occur in less than 1% of pregnancies. In some women, complications after amniocentesis may lead to a miscarriage, or loss of the pregnancy. A woman's background risk of having a miscarriage, without amniocentesis, is approximately 2-3% in her second trimester. In experienced hands, the risk for an amniocentesis-related pregnancy loss is estimated to be an additional 0.25-0.50%, or roughly one in every 200-400 pregnancies.

Much attention is often paid to the physical side effects of amniocentesis. However, it is important to also emphasize some of the emotional side effects of amniocentesis. Many of these are also applicable to other forms of prenatal diagnosis.

The offer of prenatal testing is associated with increased anxiety. This appears to be true whether a woman knew prenatal testing would be offered to her during the pregnancy or if it comes about unexpectedly, as is usually the case following abnormal screening results. Women to whom genetic amniocentesis is presented must consider the perceived benefits of testing, such as the reassurance that comes when results are normal, and compare them to the possible risks. Examples of potential risks include not only the risk of a complication after testing but also the potential burden of knowing that she will have a child with a serious disability or chronic medical condition. The nature of the child's possible diagnosis is also important. For example, could it lead to an early death, be more subtle and cause few outward signs of a problem, or be somewhere in between? There are few treatments available to correct the hundreds of known genetic disorders. Couples may wish to consider whether or not they would consider a termination of the pregnancy if a serious abnormality were detected. The definition of serious is often a matter of personal opinion. A couple's value system and family history, including that of other pregnancies and their outcomes, all influence their decision regarding amniocentesis. Ideally, a woman and her partner will have discussed at least some of these issues with each other and with either the woman's doctor or a genetic counselor prior to testing. The choice to have amniocentesis depends on many factors and should remain a personal decision.

Results

It is important to emphasize that normal results from tests performed on amniotic fluid do not necessarily guarantee the birth of a normal infant. Each couple in the general population faces a risk of roughly 3-4% of having a child with any type of congenital birth defect. Many of these will not be detected with tests performed on amniotic fluid samples obtained by amniocentesis. Babies with birth defects are often born into families with no history of genetic disorders.

Chromosome karyotyping is interpreted according to standardized nomenclature, International System of Cytogenetic Nomenclature (ISCN). The system describes the number of chromosomes, the sex chromosomes, and then any abnormalities seen in specific chromosomes using a numerical system to identify banding characteristics. Bands are numbered starting at the centromere. Standardized symbols and abbreviations describe the type of abnormality. For example, a + sign designates an extra chromosome and 47,XY, + 21 designates a male with an extra chromosome 21. The designation 46,XX,t(3:5)(q21;p15.2) means that the subject is a female with a normal number of chromosomes. There is a translocation (exchange) between chromosomes three and five involving band 21 of the long arm of number three and band 15.2 of the short arm of chromosome five.

Amniotic fluid is normally clear or slightly turbid. The concentration of AFP is reported as multiples of the median (MOM). Normal values are dependent upon the sampling time. For example, the median AFP at 15 weeks is 16.3 micrograms per mL and at 20 weeks is 8.1 micrograms per mL. Bilirubin is also time-dependent normally decreasing during gestation. The concentration should be below 0.075 mg/dL at 20 weeks and below 0.025 mg/dL at 40 weeks (full term).

An L:S ratio of 2.0 or higher correlates with fetal lung maturity. Lecithin greater than 0.10 mg/dL also correlates with fetal lung maturity.

Health care team roles

Actual samples of amniotic fluid or placenta are obtained by persons with specialized training. These are usually obstetricians or gynecologists with additional training. Sonographers assist in amniocentesis. Specially trained nurses prepare women for amniocentesis. Cytogenetic technologists, CLSp(CG) perform cell culture, karyotyping, and DNA analysis of fetal cells. Clinical laboratory scientists/medical technologists perform biochemical testing, and may also perform DNA and some molecular tests on fetal cells from the fluid. Pathologists with special training in genetics interpret results. Obstetricians or geneticists provide results to a woman or couple and discuss treatment options that may be available. Genetic counselors and other mental health professionals may provide additional support and counseling to a woman or couple after the results of amniocentesis have been returned.

Resources

BOOKS

Elias, Sherman, Simpson, Joe Leigh, and Bombard, Allan T. "Amniocentesis and Fetal Blood Sampling." In Genetic Disorders in the Fetus: Diagnosis, Prevention, and Treatment, 5th Edition, edited by Aubrey Milunsky. Baltimore: Johns Hopkins University Press, 1998, 53-82.

KEY TERMS

Alpha fetoprotein— A protein found in amniotic fluid that is commonly used to screen for certain structural birth defects.

Amnion— A thin, transparent membrane that holds the fetus in the amniotic fluid. The amniotic sac is sometimes called the bag of waters.

Anesthetic— Drug used to temporarily cause loss of sensation in an area of the body. An anesthetic may either be general, associated with a loss of consciousness, or local, affecting one area only without loss of consciousness. Anesthetics are administered via inhalation, topical application or needle injection.

Aneuploid— A fetus with an abnormal number of chromosomes.

Chorion— The outer membrane of the amniotic sac. Chorionic villi develop from its outer surface early in pregnancy. The villi establish a physical connection with the wall of the uterus. The chorionic villi eventually develop into the placenta.

Chromosome— A linear thread of genetic material contained within every cell. Humans have 46 chromosomes arranged into 23 distinct pairs. Each parent contributes one set of chromosomes, or 23, to a child. Changes in the total number of chromosomes, the shape and size (structure) of a chromosome or the contents of a chromosome may lead to abnormalities in the amount of genetic material. These abnormalities often lead to abnormal physical and mental development.

Conceptus— The product of conception, or the union of a sperm and egg cell at fertilization.

Cystic fibrosis— An inherited disease characterized by repeated lung infections, functional abnormalities of the pancreas, and an elevated level of salt in sweat. Individuals with cystic fibrosis require long-term aggressive medical care. Survival into adulthood is common, in part due to advances in treatment. Death, however, is frequently caused by respiratory failure. Although cystic fibrosis is more common among Caucasians, it has been reported in individuals of other races.

Down syndrome— A genetic condition characterized by moderate to severe mental retardation, a characteristic facial appearance, and, in some individuals, abnormalities of some internal organs. Down syndrome is always caused by an extra copy of chromosome 21, thus there are three rather than the normal two. For this reason, Down syndrome is also known as trisomy 21.

Fetus— The term used to describe a developing human infant from approximately the third month of pregnancy until delivery. The term embryo is used prior to the third month.

Fibroid— A non-cancerous tumor of connective tissue. It is made up of elongated, threadlike structures, or fibers, which usually grow slowly and are contained within an irregular shape. Fibroids are firm in consistency but may become painful if they start to break down or apply pressure to areas within the body. They frequently occur in the uterus and are generally left alone unless growing rapidly or causing other problems. Surgery is needed to remove fibroids.

Sickle cell anemia— An hereditary form of anemia due to abnormal sickle-shaped red blood cells. As a result, the cells cannot efficiently carry oxygen to body tissues. Common clinical features of sickle cell anemia include severe pain in the joints and abdomen, swelling of the tops of hands and feet, and fever. Sickle cell anemia is frequently found among individuals with ancestors who lived in central Africa.

Tay-Sachs disease— An inherited biochemical disease caused by lack of a specific enzyme in the body. In classical Tay-Sachs disease, previously normal children become blind and mentally handicapped, develop seizures, and rapidly decline. Death often occurs between the ages of three to five years. Tay-Sachs disease is common among individuals of eastern European Jewish background but has been reported in other ethnic groups.

Trimester— A three-month period. Human pregnancies are normally divided into three trimesters: first (conception to week 12), second (week 13 to week 24), and third (week 25 until delivery).

Uterus— A muscular, hollow organ of the female reproductive tract. The uterus contains and nourishes an embryo or fetus from the time the fertilized egg is implanted until birth.

Rapp, Rayna. Testing Women, Testing the Fetus: The Social Impact of Amniocentesis in America. New York: Routledge, 2000.

Shannon, Joyce B. "Amniocentesis and chorionic villus sampling (CVS)." In Health Reference Series, Medical Tests Sourcebook. Joyce B. Shannon, Editor. Detroit, MI, Omniographics, Inc., 1999, 517-522.

Wexler, Keith and Wexler, Laurie. The ABC's of Prenatal Diagnosis. Denver, CO: Genassist Inc., 1994.

PERIODICALS

Bahado-Singh, R.O., Oz, U., Hsu, C.D., Deren, O., Copel, J.A., Mahoney, M.J. "Ratio of nuchal thickness to humerus length for Down syndrome detection." American Journal of Obstetrics and Gynecology 184, no. 6. (2001): 1284-1288.

Brambati, B., Tului, L., Guercilena, S., Alberti, E. "Outcome of first-trimester chorionic villus sampling for genetic investigation in multiple pregnancy." Ultrasound in Obstetrics and Gynecology 17, no. 3 (2001): 209-216.

Canadian Early and Mid-trimester Amniocentesis Trial (CEMAT) Group. "Randomized trial to assess the safety and fetal outcome of early and mid-trimester amniocentesis." Lancet 351 (January 24, 1998): 242-247.

Cederholm, M., Sjoden, P.O., Axelsson, O. "Psychological distress before and after prenatal invasive karyotyping." Acta Obstetrics and Gynecology of Scandinavia 80, no. 6 (2001): 539-545.

Minola, E., Maccabruni, A., Pacati, I., Martinetti, M. "Amniocentesis as a possible risk factor for mother-to-infant transmission of hepatitis C virus." Hepatology 33, no. 5 (2001): 1341-1342.

Stoler, J.M., Bromley, B., Castro, M.A., Cole, W.G., Florer, J., Wenstrup, R.J. "Separation of amniotic membranes after amniocentesis in an individual with the classic form of EDS and haploinsufficiency for COL5A1 expression." American Journal of Medical Genetics 101, no. 2 (2001): 174-177.

Tercyak, K.P., Johnson, S.B., Roberts, S.F., Cruz, A.C. "Psychological response to prenatal genetic counseling and amniocentesis." Patient Education and Counseling 43, no. 1 (2001): 73-84.

ORGANIZATIONS

American Association for Clinical Chemistry, 2101 L Street, NW Suite 202, Washington, DC 20037-1558. (800) 892-1400 or (202) 857-0717. Fax: (202) 887-5093. [email protected].

American Board of Obstetrics and Gynecology, 2915 Vine Street Suite 300, Dallas TX. 75204. (214) 871-1619. Fax: (214) 871-1943. 〈http://www.abog.org/〉. [email protected].

American College of Obstetricians and Gynecologists, 409 12th St., S.W., PO Box 96920, Washington, D.C. 20090-6920. 〈http://www.acog.org/〉.

American Infertility Association, 666 Fifth Avenue, Suite 278, New York, NY 10103. (718) 621-5083. 〈http://www.americaninfertility.org/〉. [email protected].

American Society for Reproductive Medicine. 1209 Montgomery Highway, Birmingham, AL 35216-2809. (205) 978-5000. 〈http://www.asrm.com〉.

OTHER

American Academy of Family Physicians. 〈http://family-doctor.org/handouts/144.html〉.

Medical College of Pennsylvania Hahnemann University. 〈http://www.mcphu.edu/institutes/iwh/whe/briefs/amnio.htm〉.

National Library of Medicine. 〈http://www.nlm.nih.gov/medlineplus/ency/article/003921.htm〉.

Neonatology on the Web. 〈http://www.neonatology.org/syllabus/amniocentesis.html〉.

Royal College of Obstetricians and Gynecologists. 〈http://www.rcog.org.uk/guidelines/amniocentesis.html〉.

Stanford University. 〈http://www.stanford.edu/∼holbrook/Amniocentesis.html〉.

University of Michigan College of Medicine. 〈http://www.med.umich.edu/1libr/womens/ob09.htm〉.

Amniocentesis

Definition

Amniocentesis is an optional procedure offered to women during pregnancy in order to obtain more information about a developing fetus. A doctor uses a thin, hollow needle to remove a small sample of amniotic fluid from around the developing baby. An ultrasound exam is usually performed at the same time to help guide the needle. The fluid sample is used to look for specific types of medical problems in the fetus. Tests done on amniotic fluid obtained by amniocentesis cannot evaluate the fetus for every potential kind of problem. The information it does provide, however, is very accurate. The procedure is associated with a slightly increased chance for pregnancy loss. Women who undergo amniocentesis typically do so either to obtain reassurance about fetal well-being or, if the results are abnormal, to plan for the remainder of their prenatal care.

Description

Amniocentesis is the most common invasive prenatal diagnosis technique offered to pregnant women. A sample of amniotic fluid can be used to detect chromosomal abnormalities in a fetus, certain other types of congenital disorders, or other medical indicators. Its safety and accuracy are well-established, and it is generally considered the "gold standard" by which other prenatal diagnosis techniques are measured.

The word amniocentesis is derived from the Greek words, amnion and kentesis, meaning "lamb" and "puncture," respectively. In order to perform the procedure, a doctor inserts a thin needle into the mother's uterus and the amniotic sac. A continuous ultrasound evaluation is typically used so that the doctor can avoid touching both the baby and the umbilical cord with the needle. The amniotic sac is made up of two membranes: the inner amnion and the outer chorion. The amnion and chorion both develop from the fertilized egg. They are initially separate but begin to fuse early in pregnancy. This fusion is usually completed by approximately the fourteenth to fifteenth week of pregnancy.

Amniocentesis is usually performed in the second trimester, usually during weeks 16–18 (mid-trimester). The amniotic sac holds the fetus suspended within the amniotic fluid, an almost colorless fluid that protects the fetus from harm, helps maintain a consistent temperature, and prevents the fetus, or parts of it, from becoming attached to the amnion. The amniotic fluid is produced and absorbed by the fetus throughout pregnancy. Fetal cells, primarily derived from the skin, digestive system, and urinary tract, are suspended within the fluid. A smaller number of cells from the amnion and placenta are also present. Finally, the fetus produces a number of different chemical substances that also pass into the amniotic fluid. These substances may be used, in some higher-risk pregnancies, either to assess fetal lung maturity or to determine if the fetus has a viral infection. In the second trimester of pregnancy, one particular protein, called alpha-fetoprotein, is commonly used to screen for certain structural birth defects.

It is possible to perform amniocentesis in a twin pregnancy. Amniocentesis in some higher-order pregnancies, such as triplets, has also been reported. In a multiple pregnancy, it is important to ensure that a separate sample of amniotic fluid is obtained from each fetus. To accomplish this, a doctor injects a small amount of harmless blue dye into the amniotic sac of the first baby after a sample has been withdrawn. The dye will temporarily tinge the fluid blue-green. A second needle is inserted into the next amniotic sac with ultrasound guidance. If the fluid withdrawn is pale yellow, a sample from the next fetus has been successfully obtained. In the case of monoamniotic (in one amniotic sac) twins or triplets, the genetic material in each fetus is identical, so only one sample needs to be taken.

Indications for amniocentesis

Amniocentesis has been considered a standard of obstetrical care since the 1970s. It is not, however, offered to all pregnant women. The American College of Obstetricians and Gynecologists (ACOG) recommends that amniocentesis be offered to all expectant mothers age 35 and older. This age cut-off has been selected because advancing maternal age is associated with an increasing risk of having a baby with a numerical chromosome abnormality. At age 35, this risk is approximately equivalent to the risk of pregnancy loss associated with amniocentesis.

A person normally has a total of 46 chromosomes in each cell of his or her body, with the exception of sperm or egg cells, which each have only 23. As women get older, there is an increased risk of producing an egg cell with an extra chromosome. This leads to an egg cell with 24 chromosomes rather than the normal 23. Pregnancies with an abnormal number of chromosomes are referred to as aneuploid. Aneuploidy results in a conceptus (product of conception) with either too much or too little genetic material. This, in turn, leads to abnormal development. Common effects of aneuploidy include an increased risk for pregnancy loss or, in live borns, for mental retardation and physical abnormalities.

Down syndrome is the most common form of aneuploidy in live born infants, occurring in approximately one in 800 births, regardless of maternal age. In women who are 35 years old, the risk of having a child with Down syndrome is higher, or roughly one in 385 at delivery. It is important to realize that Down syndrome is not the only chromosome abnormality that may occur. Other numerical abnormalities are possible, yielding genetic conditions that may be either more or less severe than Down syndrome. Thus, a woman is often given a risk, based solely on her age, of having a child with any type of chromosome abnormality. At age 35, this total risk is approximately one in 200. By age 40, this risk has increased to one in 65, and, at age 45, this risk is one in 20. These numbers reflect the risk at the time of delivery.

Women younger than 35 years may also have children with chromosomal or other genetic disorders . Therefore, other indications for amniocentesis or other forms of prenatal diagnosis include a family history of, or a previous child with, a known genetic condition; abnormal prenatal screening results, such as ultrasound or a blood test; or one parent with a previously identified structural chromosome rearrangement. All of the above may make it more likely for a couple to have a child with a genetic condition.

Side effects

Women who have had an amniocentesis often describe it as uncomfortable, involving some mild pressure or pain as the needle is inserted. Fewer women describe it as extremely painful. A local anesthetic may be used to numb the upper layer of the mother's skin prior to testing. This medicine has no effect on the fetus, but may help the mother feel more comfortable during the procedure. An experienced physician can, on average, perform amniocentesis in approximately one to two minutes.

Common complaints after amniocentesis include mild abdominal tenderness at the site of needle insertion or mild cramping. These usually go away within one to two days. More serious complications are significantly less common but include leakage of amniotic fluid, vaginal bleeding, or uterine infection. These complications are estimated to occur in fewer than 1% of pregnancies. In some women, complications after amniocentesis may lead to a miscarriage, or loss of the pregnancy. A woman's background risk of having a miscarriage, without amniocentesis, is approximately 2–3% in her second trimester. When performed by an experienced physician or technician, the risk for an amniocentesis-related pregnancy loss is estimated to be an additional 0.25%–0.50%, or roughly one in every 200–400 pregnancies.

Much attention is often paid to the physical side effects of amniocentesis. However, it is important to also emphasize some of the emotional side effects of amniocentesis. Many of these are applicable to other forms of prenatal diagnosis.

The offer of prenatal testing is associated with increased anxiety. This appears to be true whether a woman knew prenatal testing would be offered to her during the pregnancy or if it comes about unexpectedly, as is usually the case following abnormal screening results. Women to whom genetic amniocentesis is presented must consider the perceived benefits of testing, such as the reassurance that comes when results are normal, and compare them to the possible risks. Potential risks include not only complications after testing but also learning of having a child with a serious disability or chronic medical condition. The nature of the child's possible diagnosis is also important. For example, could it lead to an early death, be more subtle and cause few outward signs of a problem, or be somewhere in between? There are few treatments available to correct the hundreds of genetic disorders so far described. Couples may consider whether or not they would consider early termination of the pregnancy if a serious abnormality were detected. The definition of "serious" is often a matter of personal opinion. A couple's value system and family history, including that of other pregnancies and their outcomes, all influence their decision regarding amniocentesis. Ideally, a woman and her partner will have discussed at least some of these issues with each other and with either the woman's doctor or a genetic counselor prior to testing. The choice to have amniocentesis depends on many factors and should remain a personal decision.

Results

Genetic testing is available on amniotic fluid obtained by amniocentesis. The most common test result is a complete analysis of the fetal chromosomes. After a sample of amniotic fluid is obtained, the genetic laboratory isolates the cells, referred to as amniocytes, out of the fluid. The cells are placed into two or more containers filled with liquid nutrients, establishing different cultures in which the cells will continue to grow. The cells are cultured anywhere between one to two weeks before the actual analysis begins. This is done in order to synchronize the growth of the cells within a culture. Also, chromosomes are only microscopically visible at a specific point during cell division.

Once there appears to be an adequate number of cells to study, the cultures are harvested. Harvesting prevents additional cell growth and stops the cells at whatever point they were in their division process. A careful study of the total number and structure of the chromosomes within the cells may now be performed. Typically, chromosome results are available within 7–14 days after amniocentesis. Results may be delayed by slow-growing cultures. This rarely reflects an abnormal result but does extend the time until final results are ready.

Many laboratories are beginning to incorporate a special technique called fluorescence in situ hybridization (FISH) into their chromosome studies. This adjunct testing provides limited information about certain chromosomes within one to two days after amniocentesis. It does not replace a complete chromosome study using amniocyte cultures. In fact, FISH results are often reported as preliminary, pending confirmation by cultured results. They can, however, be very useful, particularly when there is already a high level of suspicion of a fetal chromosome abnormality.

FISH is performed using a small sample of uncultured amniotic fluid cells. Special molecular tags for particular chromosomes are used. These tags attach themselves to the chromosome. Under specific laboratory conditions, they can be made to "light up" or fluoresce. Their signals can then be counted using a special kind of microscope. FISH is most often used to quickly identify a change in the number of chromosomes from pairs 13, 18, 21, and the two sex chromosomes, X and Y. Abnormalities of these chromosomes account for nearly 95% of all chromosomal abnormalities. Other chromosomal abnormalities will be missed since FISH cannot identify structural rearrangements of the chromosomes or abnormalities involving other pairs. A full chromosome evaluation on cultured cells is a necessary follow-up to interphase FISH results.

A sample of amniotic fluid may be used to measure alpha-fetoprotein (AFP). AFP is a protein made by the fetal liver. It passes out of the fetus and enters both the amniotic fluid and the mother's blood. Screening for open neural tube defects, abnormal openings in the fetal head or spinal cord, or ventral wall defects, openings along the belly wall, can be done by measuring AFP during the fifteenth to twentieth weeks of pregnancy. AFP levels normally show a gradual increase during this time. An unusually high level of serum AFP does not necessarily indicate a problem with fetal development, but is cause for some concern. A high AFP level in amniotic fluid will detect up to 98% of all openings on the fetal body that are not covered by skin. Further studies may be suggested if the AFP is high. Most initial AFP results are available within two to three days after amniocentesis.

Finally, amniotic fluid samples obtained by amniocentesis may also be used for more specialized genetic studies, such as biochemical or DNA testing. Both often require cell cultures and additional time to complete. These studies are not done on every sample. Rather, they are offered to those couples who, based on their family history or other information, are at increased risk of having a child with a single gene , or Mendelian, disorder. Hundreds of such disorders have been described. Examples include Tay-Sachs disease , cystic fibrosis , and sickle cell anemia . If biochemical or DNA studies are performed, all of the results may not be ready until three to four weeks after testing, although for each patient, the waiting time may be slightly different.

It is important to emphasize that normal results from tests done on amniotic fluid do not necessarily guarantee the birth of a normal infant. Each couple in the general population faces a risk of roughly 3–4% of having a child with any type of congenital birth defect. Many of these will not be detected with tests done on amniotic fluid samples obtained by amniocentesis. Babies with birth defects are often born into families with no history of genetic disorders.

Chorionic villus sampling

Mid-trimester amniocentesis has been available for nearly thirty years. Chorionic villus sampling (CVS) has been available in the United States since the 1980s. CVS is usually performed between ten to twelve weeks of pregnancy. It involves the removal of a small sample of the developing placenta, or chorionic villi. It has been an attractive alternative to amniocentesis, particularly for those women who desire both testing and results earlier in their pregnancies. Some of the benefits of earlier testing include reassurance sooner in pregnancy and fewer physical complications following first trimester pregnancy termination, for those couples who choose this option after testing. CVS is, however, associated with a higher risk of miscarriage than mid-trimester amniocentesis. At experienced centers, this risk is approximately 1% (or, 1 in 100).

Early amniocentesis

Early amniocentesis is performed before the thirteenth completed week of pregnancy. It has been considered experimental for many years. The results of the largest early amniocentesis trial, published in 1998, have caused physicians worldwide to reconsider the benefit and risks of this procedure.

The Canadian early and mid-trimester amniocentesis trial (CEMAT) is the largest multi-center, randomized clinical trial of early amniocentesis to date. The purpose of the trial was to examine and compare the safety and accuracy of early (EA) versus mid-trimester amniocentesis (MTA). In order to accomplish this, 4,374 pregnant women were identified and enrolled in the study. Ultrasound was performed in the first trimester to confirm the gestational age of all pregnancies. Computer randomization was used to evenly divide the women into either the EA or MTA groups. Ultimately, 1,916 women underwent EA and 1,775 women had MTA. Follow-up was obtained on nearly all pregnancies. Two striking conclusions were reached: EA is associated with an increased incidence of clubfoot and an increased risk of procedure-related pregnancy loss.

Clubfoot, also referred to as talipes equinovarus, occurs in approximately one in 1,000 live births (0.1%) in the general population. It may involve either one foot (unilateral) or both feet (bilateral). Males are affected slightly more often than females. There are several proposed mechanisms by which clubfoot could occur: due to the interaction of several genes during development, as a direct consequence of environmental factors, such as an abnormal position in the uterus, or as a physical component of a single gene disorder. Any such disorder would be expected to also cause other abnormalities.

Overall, the CEMAT study found an incidence of clubfoot in the EA group of 1.3% (29 infants). None of the affected infants had other abnormalities. This is nearly ten times higher than the risk in the general population. The frequency of clubfoot in the MTA group was the same as in the general population (0.1%). Prior studies of mid-trimester amniocentesis did not reveal an increased frequency of infants with clubfoot or other birth defects.

Clubfoot was more common when testing was performed during the eleventh, rather than the twelfth, week of pregnancy. This suggests that there may be a specific window sometime in the eleventh to twelfth weeks during which the fetus may be particularly vulnerable to developing clubfoot. It is possible that EA causes a temporary, but still significant, loss of amniotic fluid. This loss may go unrecognized. However, it could, in turn, affect the flow of blood to the foot or cause direct pressure on the developing limb, either of which could lead to clubfoot. It is difficult to know which potential mechanism could be correct since the number of affected infants born after EA is relatively small.

Of note, a separate, much smaller, study also demonstrated an increased incidence of clubfoot (1.7%) among the set of women who underwent EA. The study consisted of patients randomized between EA and CVS and examined the risk of miscarriage after EA. Enrollment in the study was stopped once the association between EA and clubfoot was identified. There were no birth defects identified after CVS.

An additional concern recognized from CEMAT was a higher rate of miscarriage after EA. A procedure-related loss was defined as one that occurred either shortly after the testing or before twenty weeks of pregnancy. Fifty-five women (2.5%) experienced a miscarriage after EA. In contrast, miscarriage occurred in seventeen (0.8%) of the MTA patients. An increased rate of loss appeared to more often follow technically challenging procedures. Difficult procedures included those pregnancies in which bleeding occurred prior to amniocentesis or in which uterine fibroids were present. Tenting of the membranes also made early amniocentesis difficult. Tenting occurs when the amnion and chorion are not yet completely fused, as is true for the majority of first trimester pregnancies. The separation between the membranes makes insertion of the amniocentesis needle more difficult.

In the absence of a difficult EA procedure, a higher rate of loss was also observed among those pregnancies in which the mother experienced obvious leakage of amniotic fluid or vaginal bleeding after testing. The level of physician experience with EA did not influence the rate of loss.

Finally, EA was also linked to an increased number of laboratory culture failures (no growth of cells and no results) compared to MTA. The total waiting time for results was slightly longer in the EA group. This is not entirely a surprise, since a smaller amount of fluid is obtained when EA is performed. Hence, there are fewer cells, and culturing takes longer.

Demographics

According to the National Center for Health Statistics (NCHS), 112,776 amniocentesis procedures were performed in the United States in 1998, the most recent year for which data is available. The annual birth rate that year was approximately 3.9 million infants. Thus, approximately 3% of pregnant women in the United States had this procedure performed. It is likely that this is an underestimate, however. The NCHS obtains information from birth certificates registered in each state and the District of Columbia. Although almost all deliveries are registered in the United States, records are still submitted with incomplete information. It is also not possible to know how many amniocentesis procedures were performed for genetic testing, as compared to other indications, as this information is not requested.

Summary

Amniocentesis is a reliable procedure for prenatal diagnosis in the second trimester of pregnancy. It is primarily offered to pregnant women who are at increased risk, based on their age, family history, or other factor, of having a child with a genetic condition. Amniocentesis provides accurate information about fetal chromosomes or the likelihood of certain physical abnormalities. Additional specialized studies may be performed on an as-needed basis. Despite these benefits, amniocentesis is associated with a slightly increased chance of pregnancy loss. Each woman should discuss the potential risks and benefits of amniocentesis with a doctor or genetic counselor to make a decision about whether or not she has this testing. Early amniocentesis, or procedures performed before the thirteenth week of pregnancy, has been associated with an increased risk of clubfoot and of procedure-related pregnancy loss.

Resources

BOOKS

"Amniocentesis and chorionic villus sampling (CVS)." In Medical Tests Sourcebook. 1st ed. Health Reference Series, edited by Joyce Brennfleck Shannon. Detroit: Omniographics Inc., 1999, pp. 517–522.

Elias, Sherman, Joe Leigh Simpson, and Allan T. Bombard. "Amniocentesis and Fetal Blood Sampling." In Genetic Disorders and the Fetus: Diagnosis, Prevention, and Treatment. 4th ed. Edited by Aubrey Milunsky. Baltimore: The Johns Hopkins University Press, 1998, pp. 53–82.

PERIODICALS

The Canadian Early and Mid-trimester Amniocentesis Trial (CEMAT) Group. "Randomized trial to assess the safety and fetal outcome of early and mid-trimester amniocentesis." Lancet 351 (January 24, 1998): 242–247.

Farrell, Sandra A., A.M. Summers, Louis Dallaire, Joel Singer, JoAnn M. Johnson, and R. Douglas Wilson, members of CEMAT. "Club foot, an adverse outcome of early amniocentesis: disruption or deformation?" Journal Medical Genetics 36, no. 11 (November 1999): 843–846.

ORGANIZATIONS

American College of Obstetricians and Gynecologists. PO Box 96920, 409 12th St. SW, Washington, DC 20090-6920. <http://www.acog.org>.

National Center for Health Statistics. Division of Data Services, 6525 Belcrest Rd., Hyattsville, MD 20782-2003. <http://www.cdc.gov/nchs>.

WEBSITES

"Amniocentesis." http://www.medicinenet.com/script/main/Art.asp?li=MN1&ArticleKey=268.

"Amniocentesis." <http://www.modimes.org/HealthLibrary2/factsheets/Amniocentesis.htm>.

"Prenatal diagnosis: Amniocentesis and CVS." <http://www.familydoctor.org/handouts/144.html>.

Terri A. Knutel, MS, CGC

Amniocentesis

Definition

Amniocentesis is an optional procedure offered to women during pregnancy in order to obtain more information about a developing fetus. A doctor uses a thin, hollow needle to remove a small sample of amniotic fluid from around the developing baby. An ultrasound exam is usually performed at the same time to help guide the needle. The fluid sample is used to look for specific types of medical problems in the fetus. Tests done on amniotic fluid obtained by amniocentesis cannot evaluate the fetus for every potential kind of problem. The information it does provide, however, is very accurate. The procedure is associated with a slightly increased chance for pregnancy loss. Women who undergo amniocentesis typically do so either to obtain reassurance about fetal well-being or, if the results are abnormal, to plan for the remainder of their prenatal care.

Description

Amniocentesis is the most common invasive prenatal diagnosis technique offered to pregnant women. A sample of amniotic fluid can be used to detect chromosomal abnormalities in a fetus, certain other types of congenital disorders, or other medical indicators. Its safety and accuracy are well-established, and it is generally considered the "gold standard" by which other prenatal diagnosis techniques are measured.

The word amniocentesis is derived from the Greek words, amnion and kentesis, meaning "lamb" and "puncture," respectively. In order to perform the procedure, a doctor inserts a thin needle into the mother's uterus and the amniotic sac. A continuous ultrasound evaluation is typically used so that the doctor can avoid touching both the baby and the umbilical cord with the needle. The amniotic sac is made up of two membranes: the inner amnion and the outer chorion. The amnion and chorion both develop from the fertilized egg. They are initially separate but begin to fuse early in pregnancy. This fusion is usually completed by approximately the fourteenth to fifteenth week of pregnancy.

Amniocentesis is usually performed in the second trimester, usually during weeks 16–18 (mid-trimester). The amniotic sac holds the fetus suspended within the amniotic fluid, an almost colorless fluid that protects the fetus from harm, helps maintain a consistent temperature, and prevents the fetus, or parts of it, from becoming attached to the amnion. The amniotic fluid is produced and absorbed by the fetus throughout pregnancy. Fetal cells, primarily derived from the skin, digestive system, and urinary tract, are suspended within the fluid. A smaller number of cells from the amnion and placenta are also present. Finally, the fetus produces a number of different chemical substances that also pass into the amniotic fluid. These substances may be used, in some higher-risk pregnancies, either to assess fetal lung maturity or to determine if the fetus has a viral infection. In the second trimester of pregnancy, one particular protein, called alpha-fetoprotein, is commonly used to screen for certain structural birth defects.

It is possible to perform amniocentesis in a twin pregnancy. Amniocentesis in some higher-order pregnancies, such as triplets, has also been reported. In a multiple pregnancy, it is important to ensure that a separate sample of amniotic fluid is obtained from each fetus. To accomplish this, a doctor injects a small amount of harmless blue dye into the amniotic sac of the first baby after a sample has been withdrawn. The dye will temporarily tinge the fluid blue-green. A second needle is inserted into the next amniotic sac with ultrasound guidance. If the fluid withdrawn is pale yellow, a sample from the next fetus has been successfully obtained. In the case of monoamniotic (in one amniotic sac) twins or triplets, the genetic material in each fetus is identical, so only one sample needs to be taken.

Indications for amniocentesis

Amniocentesis has been considered a standard of obstetrical care since the 1970s. It is not, however, offered to all pregnant women. The American College of Obstetricians and Gynecologists (ACOG) recommends that amniocentesis be offered to all expectant mothers age 35 and older. This age cut-off has been selected because advancing maternal age is associated with an increasing risk of having a baby with a numerical chromosome abnormality. At age 35, this risk is approximately equivalent to the risk of pregnancy loss associated with amniocentesis.

A person normally has a total of 46 chromosomes in each cell of his or her body, with the exception of sperm or egg cells, which each have only 23. As women get older, there is an increased risk of producing an egg cell with an extra chromosome. This leads to an egg cell with 24 chromosomes rather than the normal 23. Pregnancies with an abnormal number of chromosomes are referred to as aneuploid. Aneuploidy results in a conceptus (product of conception) with either too much or too little genetic material. This, in turn, leads to abnormal development. Common effects of aneuploidy include an increased risk for pregnancy loss or, in live borns, for mental retardation and physical abnormalities.

Down syndrome is the most common form of aneuploidy in live born infants, occurring in approximately 1 in 800 births, regardless of maternal age. In women who are 35 years old, the risk of having a child with Down syndrome is higher, or roughly one in 385 at delivery. It is important to realize that Down syndrome is not the only chromosome abnormality that may occur. Other numerical abnormalities are possible, yielding genetic conditions that may be either more or less severe than Down syndrome. Thus, a woman is often given a risk, based solely on her age, of having a child with any type of chromosome abnormality. At age 35, this total risk is approximately one in 200. By age 40, this risk has increased to one in 65, and, at age 45, this risk is one in 20. These numbers reflect the risk at the time of delivery.

Women younger than 35 years may also have children with chromosomal or other genetic disorders . Therefore, other indications for amniocentesis or other forms of prenatal diagnosis include a family history of, or a previous child with a known genetic condition; abnormal prenatal screening results, such as ultrasound or a blood test; or one parent with a previously identified structural chromosome rearrangement. All of the above may make it more likely for a couple to have a child with a genetic condition.

Side effects

Women who have had an amniocentesis often describe it as uncomfortable, involving some mild pressure or pain as the needle is inserted. Fewer women describe it as extremely painful. A local anesthetic may be used to numb the upper layer of the mother's skin prior to testing. This medicine has no effect on the fetus, but may help the mother feel more comfortable during the procedure. An experienced physician can, on average, perform amniocentesis in approximately one to two minutes.

Common complaints after amniocentesis include mild abdominal tenderness at the site of needle insertion or mild cramping. These usually go away within one to two days. More serious complications are significantly less common but include leakage of amniotic fluid, vaginal bleeding, or uterine infection. These complications are estimated to occur in fewer than 1% of pregnancies. In some women, complications after amniocentesis may lead to a miscarriage, or loss of the pregnancy. A woman's background risk of having a miscarriage, without amniocentesis, is approximately 2–3% in her second trimester. When performed by an experienced physician or technician, the risk for an amniocentesis-related pregnancy loss is estimated to be an additional 0.25%–0.50%, or roughly one in every 200–400 pregnancies.

Much attention is often paid to the physical side effects of amniocentesis. However, it is important to also emphasize some of the emotional side effects of amniocentesis. Many of these are applicable to other forms of prenatal diagnosis.

The offer of prenatal testing is associated with increased anxiety. This appears to be true whether a woman knew prenatal testing would be offered to her during the pregnancy or if it comes about unexpectedly, as is usually the case following abnormal screening results. Women to whom genetic amniocentesis is presented must consider the perceived benefits of testing, such as the reassurance that comes when results are normal, and compare them to the possible risks. Potential risks include not only complications after testing but also learning that the child may have a serious disability or chronic medical condition. The nature of the child's possible diagnosis is also important. For example, could it lead to an early death, be more subtle and cause few outward signs of a problem, or be somewhere in between? There are few treatments available to correct the hundreds of genetic disorders so far described. Couples may consider early termination of the pregnancy if a serious abnormality were detected. The definition of "serious" is often a matter of personal opinion. A couple's value system and family history, including that of other pregnancies and their outcomes, all influence their decision regarding amniocentesis. Ideally, a woman and her partner will have discussed at least some of these issues with each other and with either the woman's doctor or a genetic counselor prior to testing. The choice to have amniocentesis depends on many factors and should remain a personal decision.

Results

Genetic testing is available on amniotic fluid obtained by amniocentesis. The most common test result is a complete analysis of the fetal chromosomes. After a sample of amniotic fluid is obtained, the genetic laboratory isolates the cells, referred to as amniocytes, out of the fluid. The cells are placed into two or more containers filled with liquid nutrients, establishing different cultures in which the cells will continue to grow. The cells are cultured anywhere between one to two weeks before the actual analysis begins. This is done in order to synchronize the growth of the cells within a culture. Also, chromosomes are only microscopically visible at a specific point during cell division.

Once there appears to be an adequate number of cells to study, the cultures are harvested. Harvesting prevents additional cell growth and stops the cells at whatever point they were in their division process. A careful study of the total number and structure of the chromosomes within the cells may now be performed. Typically, chromosome results are available within 7–14 days after amniocentesis. Results may be delayed by slow-growing cultures. This rarely reflects an abnormal result but does extend the time until final results are ready.

Many laboratories are beginning to incorporate a special technique called fluorescence in situ hybridization (FISH) into their chromosome studies. This adjunct testing provides limited information about certain chromosomes within one to two days after amniocentesis. It does not replace a complete chromosome study using amniocyte cultures. In fact, FISH results are often reported as preliminary, pending confirmation by cultured results. They can, however, be very useful, particularly when there is already a high level of suspicion of a fetal chromosome abnormality.

FISH is performed using a small sample of uncultured amniotic fluid cells. Special molecular tags for particular chromosomes are used. These tags attach themselves to the chromosome. Under specific laboratory conditions, they can be made to "light up" or fluoresce. Their signals can then be counted using a special kind of microscope. FISH is most often used to quickly identify a change in the number of chromosomes from pairs 13, 18, 21, and the two sex chromosomes, X and Y. Abnormalities of these chromosomes account for nearly 95% of all chromosomal abnormalities. Other chromosomal abnormalities will be missed since FISH cannot identify structural rearrangements of the chromosomes or abnormalities involving other pairs. A full chromosome evaluation on cultured cells is a necessary follow-up to interphase FISH results.

A sample of amniotic fluid may be used to measure alpha-fetoprotein (AFP). AFP is a protein made by the fetal liver. It passes out of the fetus and enters both the amniotic fluid and the mother's blood. Screening for open neural tube defects , abnormal openings in the fetal head or spinal cord, or ventral wall defects, openings along the belly wall, can be done by measuring AFP during the fifteenth to twentieth weeks of pregnancy. AFP levels normally show a gradual increase during this time. An unusually high level of serum AFP does not necessarily indicate a problem with fetal development, but is cause for some concern. A high AFP level in amniotic fluid will detect up to 98% of all openings on the fetal body that are not covered by skin. Further studies may be suggested if the AFP is high. Most initial AFP results are available within two to three days after amniocentesis.

Finally, amniotic fluid samples obtained by amniocentesis may also be used for more specialized genetic studies, such as biochemical or DNA testing. Both often require cell cultures and additional time to complete. These studies are not done on every sample. Rather, they are offered to those couples who, based on their family history or other information, are at increased risk of having a child with a single gene ,or Mendelian, disorder. Hundreds of such disorders have been described. Examples include Tay-Sachs disease, cystic fibrosis , and sickle cell anemia. If biochemical or DNA studies are performed, all of the results may not be ready until three to four weeks after testing, although for each patient, the waiting time may be slightly different.

It is important to emphasize that normal results from tests done on amniotic fluid do not necessarily guarantee the birth of a normal infant. Each couple in the general population faces a risk of roughly 3–4% of having a child with any type of congenital birth defect. Many of these will not be detected with tests done on amniotic fluid samples obtained by amniocentesis. Babies with birth defects are often born into families with no history of genetic disorders.

Chorionic villus sampling

Mid-trimester amniocentesis has been available for nearly thirty years. Chorionic villus sampling (CVS) has been available in the United States since the 1980s. CVS is usually performed between ten to twelve weeks of pregnancy. It involves the removal of a small sample of the developing placenta, or chorionic villi. It has been an attractive alternative to amniocentesis, particularly for those women who desire both testing and results earlier in their pregnancies. Some of the benefits of earlier testing include reassurance sooner in pregnancy and fewer physical complications following first trimester pregnancy termination, for those couples who choose this option after testing. CVS is, however, associated with a higher risk of miscarriage than mid-trimester amniocentesis. At experienced centers, this risk is approximately 1% (or, one in 100).

Early amniocentesis

Early amniocentesis is performed before the thirteenth completed week of pregnancy. It has been considered experimental for many years. The results of the largest early amniocentesis trial, published in 1998, have caused physicians worldwide to reconsider the benefit and risks of this procedure.

The Canadian early and mid-trimester amniocentesis trial (CEMAT) is the largest multi-center, randomized clinical trial of early amniocentesis to date. The purpose of the trial was to examine and compare the safety and accuracy of early (EA) versus mid-trimester amniocentesis (MTA). In order to accomplish this, 4,374 pregnant women were identified and enrolled in the study. Ultrasound was performed in the first trimester to confirm the gestational age of all pregnancies. Computer randomization was used to evenly divide the women into either the EA or MTA groups. Ultimately, 1,916 women underwent EA and 1,775 women had MTA. Follow-up was obtained on nearly all pregnancies. Two striking conclusions were reached: EA is associated with an increased incidence of clubfoot and an increased risk of procedure-related pregnancy loss.

Clubfoot, also referred to as talipes equinovarus, occurs in approximately one in 1,000 live births (0.1%) in the general population. It may involve either one foot (unilateral) or both feet (bilateral). Males are affected slightly more often than females. There are several proposed mechanisms by which clubfoot could occur: due to the interaction of several genes during development, as a direct consequence of environmental factors, such as an abnormal position in the uterus, or as a physical component of a single gene disorder. Any such disorder would be expected to also cause other abnormalities.

Overall, the CEMAT study found an incidence of clubfoot in the EA group of 1.3% (29 infants). None of the affected infants had other abnormalities. This is nearly ten times higher than the risk in the general population. The frequency of clubfoot in the MTA group was the same as in the general population (0.1%). Prior studies of mid-trimester amniocentesis did not reveal an increased frequency of infants with clubfoot or other birth defects.

Clubfoot was more common when testing was performed during the eleventh, rather than the twelfth, week of pregnancy. This suggests that there may be a specific window sometime in the eleventh to twelfth weeks during which the fetus may be particularly vulnerable to developing clubfoot. It is possible that EA causes a temporary, but still significant, loss of amniotic fluid. This loss may go unrecognized. However, it could, in turn, affect the flow of blood to the foot or cause direct pressure on the developing limb, either of which could lead to clubfoot. It is difficult to know which potential mechanism could be correct since the number of affected infants born after EA is relatively small.

Of note, a separate, much smaller, study also demonstrated an increased incidence of clubfoot (1.7%) among the set of women who underwent EA. The study consisted of patients randomized between EA and CVS and examined the risk of miscarriage after EA. Enrollment in the study was stopped once the association between EA and clubfoot was identified. There were no birth defects identified after CVS.

An additional concern recognized from CEMAT was a higher rate of miscarriage after EA. A procedure-related loss was defined as one that occurred either shortly after the testing or before twenty weeks of pregnancy. Fifty-five women (2.5%) experienced a miscarriage after EA. In contrast, miscarriage occurred in seventeen (0.8%) of the MTA patients. An increased rate of loss appeared to more often follow technically challenging procedures. Difficult procedures included those pregnancies in which bleeding occurred prior to amniocentesis or in which uterine fibroids were present. Tenting of the membranes also made early amniocentesis difficult. Tenting occurs when the amnion and chorion are not yet completely fused, as is

true for the majority of first trimester pregnancies. The separation between the membranes makes insertion of the amniocentesis needle more difficult.

In the absence of a difficult EA procedure, a higher rate of loss was also observed among those pregnancies in which the mother experienced obvious leakage of amniotic fluid or vaginal bleeding after testing. The level of physician experience with EA did not influence the rate of loss.

Finally, EA was also linked to an increased number of laboratory culture failures (no growth of cells and no results) compared to MTA. The total waiting time for results was slightly longer in the EA group. This is not entirely a surprise, since a smaller amount of fluid is obtained when EA is performed. Hence, there are fewer cells, and culturing takes longer.

Demographics

According to the National Center for Health Statistics (NCHS), 112,776 amniocentesis procedures were performed in the United States in 1998, the most recent year for which data is available. The annual birth rate that year was approximately 3.9 million infants. Thus, approximately 3% of pregnant women in the United States had this procedure performed. It is likely that this is an underestimate, however. The NCHS obtains information from birth certificates registered in each state and the District of Columbia. Although almost all deliveries are registered in the United States, records are still submitted with incomplete information. It is also not possible to know how many amniocentesis procedures were performed for genetic testing, as compared to other indications, as this information is not requested.

Summary

Amniocentesis is a reliable procedure for prenatal diagnosis in the second trimester of pregnancy. It is primarily offered to pregnant women who are at increased risk, based on their age, family history, or other factor, of having a child with a genetic condition. Amniocentesis provides accurate information about fetal chromosomes or the likelihood of certain physical abnormalities. Additional specialized studies may be performed on an as-needed basis. Despite these benefits, amniocentesis is associated with a slightly increased chance of pregnancy loss. Each woman should discuss the potential risks and benefits of amniocentesis with a doctor or genetic counselor to make a decision about whether or not she has this testing. Early amniocentesis, or procedures performed before the thirteenth week of pregnancy, has been associated with an increased risk of clubfoot and of procedure-related pregnancy loss.

Resources

BOOKS

"Amniocentesis and chorionic villus sampling (CVS)." In Medical Tests Sourcebook. 1st ed. Health Reference Series, edited by Joyce Brennfleck Shannon, Detroit: Omniographics Inc., 1999, pp. 517–522.

Elias, Sherman, Joe Leigh Simpson, and Allan T. Bombard. "Amniocentesis and Fetal Blood Sampling." In Genetic Disorders and the Fetus: Diagnosis, Prevention, and Treatment. 4th ed. Edited by Aubrey Milunsky, Baltimore: The Johns Hopkins University Press, 1998, pp. 53–82.

PERIODICALS

The Canadian Early and Mid-trimester Amniocentesis Trial (CEMAT) Group. "Randomized trial to assess the safety and fetal outcome of early and mid-trimester amniocentesis." Lancet 351 (January 24, 1998): 242–247.

Farrell, Sandra A., A.M. Summers, Louis Dallaire, Joel Singer, Jo Ann M. Johnson, and R. Douglas Wilson, members of CEMAT. "Club foot, an adverse outcome of early amniocentesis: disruption or deformation?" Journal Medical Genetics 36, no. 11 (November 1999): 843–846.

ORGANIZATIONS

American College of Obstetricians and Gynecologists. PO Box 96920, 409 12th St. SW, Washington, DC 20090-6920. <http://www.acog.org>.

National Center for Health Statistics. Division of Data Services, 6525 Belcrest Rd., Hyattsville, MD 20782-2003. <http://www.cdc.gov/nchs>.

WEBSITES

"Amniocentesis." http://www.medicinenet.com/script/main/Art.asp?li=MN1&ArticleKey=268.

"Amniocentesis." <http://www.modimes.org/HealthLibrary2/factsheets/Amniocentesis.htm>.

"Prenatal diagnosis: Amniocentesis and CVS." <http://www.familydoctor.org/handouts/144.html>.

Terri A. Knutel, MS, CGC

Amniocentesis

Definition

Amniocentesis is a procedure used to diagnose fetal defects in the early second trimester of pregnancy. A sample of the amniotic fluid, which surrounds a fetus in the womb, is collected through a pregnant woman's abdomen using a needle and syringe. Tests performed on fetal cells found in the sample can reveal the presence of many types of genetic disorders, thus allowing doctors and prospective parents to make important decisions about early treatment and intervention.

Purpose

Since the mid-1970s, amniocentesis has been used routinely to test for Down syndrome, by far the most common, nonhereditary, genetic birth defect, afflicting about one in every 1,000 babies. By 1997, approximately 800 different diagnostic tests were available, most of them for hereditary genetic disorders such as Tay-Sachs disease, sickle cell anemia, hemophilia, muscular dystrophy and cystic fibrosis.

Amniocentesis, often called amnio, is recommended for women who will be older than 35 on their due-date. It is also recommended for women who have already borne children with birth defects, or when either of the parents has a family history of a birth defect for which a diagnostic test is available. Another reason for the procedure is to confirm indications of Down syndrome and certain other defects which may have shown up previously during routine maternal blood screening.

The risk of bearing a child with a nonhereditary genetic defect such as Down syndrome is directly related to a woman's age—the older the woman, the greater the risk. Thirty-five is the recommended age to begin amnio testing because that is the age at which the risk of carrying a fetus with such a defect roughly equals the risk of miscarriage caused by the procedure-about one in 200. At age 25, the risk of giving birth to a child with this type of defect is about one in 1,400; by age 45 it increases to about one in 20. Nearly half of all pregnant women over 35 in the United States undergo amniocentesis and many younger women also decide to have the procedure. Notably, some 75% of all Down syndrome infants born in the United States each year are to women younger than 35.

One of the most common reasons for performing amniocentesis is an abnormal alpha-fetoprotein (AFP) test. Alpha-fetoprotein is a protein produced by the fetus and present in the mother's blood. A simple blood screening, usually conducted around the 15th week of pregnancy, can determine the AFP levels in the mother's blood. Levels that are too high or too low may signal possible fetal defects. Because this test has a high false-positive rate, another test such as amnio is recommended whenever the AFP levels fall outside the normal range.

Amniocentesis is generally performed during the 16th week of pregnancy, with results usually available within three weeks. It is possible to perform an amnio as early as the 11th week but this is not usually recommended because there appears to be an increased risk of miscarriage when done at this time. The advantage of early amnio and speedy results lies in the extra time for decision making if a problem is detected. Potential treatment of the fetus can begin earlier. Important, also, is the fact that elective abortions are safer and less controversial the earlier they are performed.

Precautions

As an invasive surgical procedure, amnio poses a real, although small, risk to the health of a fetus. Parents must weigh the potential value of the knowledge gained, or indeed the reassurance that all is well, against the small risk of damaging what is in all probability a normal fetus. The serious emotional and ethical dilemmas that adverse test results can bring must also be considered. The decision to undergo amnio is always a matter of personal choice.

KEY TERMS

Alpha-fetoprotein (AFP)— A protein normally produced by the liver of a fetus and detectable in maternal blood samples. AFP screening measures the amount of alpha-fetoprotein in the blood. Levels outside the norm may indicate fetal defects.

Anencephaly— A hereditary defect resulting in the partial to complete absence of a brain and spinal cord. It is fatal.

Chorionic villus sampling (CVS)— A procedure similar to amniocentesis, except that cells are taken from the chorionic membrane for testing. These cells, called chorionic villus cells, eventually become the placenta. The samples are collected either through the abdomen, as in amnio, or through the vagina. CVS can be done earlier in the pregnancy than amnio, but carries a somewhat higher risk.

Chromosome— Chromosomes are the strands of genetic material in a cell that occur in nearly identical pairs. Normal human cells contain 23 chromosome pairs-one in each pair inherited from the mother, and one from the father. Every human cell contains the exact same set of chromosomes.

Down syndrome— The most prevalent of a class of genetic defects known as trisomies, in which cells contain three copies of certain chromosomes rather than the usual two. Down syndrome, or trisomy 21, usually results from three copies of chromosome 21.

Genetic— The term refers to genes, the basic units of biological heredity, which are contained on the chromosomes, and contain chemical instructions which direct the development and functioning of an individual.

Hereditary— Something which is inherited-passed down from parents to offspring. In biology and medicine, the word pertains to inherited genetic characteristics.

Maternal blood screening— Maternal blood screening is normally done early in pregnancy to test for a variety of conditions. Abnormal amounts of certain proteins in a pregnant woman's blood raise the probability of fetal defects. Amniocentesis is recommended if such a probability occurs.

Tay-Sachs disease— An inherited disease prevalent among the Ashkenazi Jewish population of the United States. Infants with the disease are unable to process a certain type of fat which accumulates in nerve and brain cells, causing mental and physical retardation, and death by age four.

Ultrasound— A technique which uses high-frequency sound waves to create a visual image (a sonogram) of soft tissues. The technique is routinely used in prenatal care and diagnosis.

Description

The word amniocentesis literally means "puncture of the amnion," the thin-walled sac of fluid in which a developing fetus is suspended during pregnancy. During the sampling procedure, the obstetrician inserts a very fine needle through the woman's abdomen into the uterus and amniotic sac and withdraws approximately one ounce of amniotic fluid for testing. The relatively painless procedure is performed on an outpatient basis, sometimes using local anesthesia.

The physician uses ultrasound images to guide needle placement and collect the sample, thereby minimizing the risk of fetal injury and the need for repeated needle insertions. Once the sample is collected, the woman can return home after a brief observation period. She may be instructed to rest for the first 24 hours and to avoid heavy lifting for two days.

The sample of amniotic fluid is sent to a laboratory where fetal cells contained in the fluid are isolated and grown in order to provide enough genetic material for testing. This takes about seven to 14 days. The material is then extracted and treated so that visual examination for defects can be made. For some disorders, like Tay-Sachs, the simple presence of a telltale chemical compound in the amniotic fluid is enough to confirm a diagnosis. Depending on the specific tests ordered, and the skill of the lab conducting them, all the results are available between one and four weeks after the sample is taken.

Cost of the procedure depends on the doctor, the lab, and the tests ordered. Most insurers provide coverage for women over 35, as a follow-up to positive maternal blood screening results, and when genetic disorders run in the family.

An alternative to amnio, now in general use, is chorionic villus sampling, or CVS, which can be performed as early as the eighth week of pregnancy. While this allows for the possibility of a first trimester abortion, if warranted, CVS is apparently also riskier and is more expensive. The most promising area of new research in prenatal testing involves expanding the scope and accuracy of maternal blood screening as this poses no risk to the fetus.

Preparation

It is important for a woman to fully understand the procedure and to feel confident in the obstetrician performing it. Evidence suggests that a physician's experience with the procedure reduces the chance of mishap. Almost all obstetricians are experienced in performing amniocentesis. The patient should feel free to ask questions and seek emotional support before, during and after the amnio is performed.

Aftercare

Necessary aftercare falls into two categories, physical and emotional.

Physical aftercare

During and immediately following the sampling procedure, a woman may experience dizziness, nausea, a rapid heartbeat, and cramping. Once past these immediate hurdles, the physician will send the woman home with instructions to rest and to report any complications requiring immediate treatment, including:

• vaginal bleeding. The appearance of blood could signal a problem.
• premature labor. Unusual abdominal pain and/or cramping may indicate the onset of premature labor. Mild cramping for the first day or two following the procedure is normal.
• signs of infection. Leaking of amniotic fluid or unusual vaginal discharge, and fever could signal the onset of infection.

Emotional aftercare

Once the procedure has been safely completed, the anxiety of waiting for the test results can prove to be the worst part of the process. A woman should seek and receive emotional support from family and friends, as well as from her obstetrician and family doctor. Professional counseling may also prove necessary, particularly if a fetal defect is discovered.

Risks

Most of the risks and short-term side effects associated with amniocentesis relate to the sampling procedure and have been discussed above. A successful amnio sampling results in no long-term side effects. Risks include:

• maternal/fetal hemorrhaging. While spotting in pregnancy is fairly common, bleeding following amnio should always be investigated.
• infection. Infection, although rare, can occur after amniocentesis. An unchecked infection can lead to severe complications.
• fetal injury. A very slight risk of injury to the fetus resulting from contact with the amnio needle does exist.
• miscarriage. The rate of miscarriage occurring during standard, second trimester amnio appears to be approximately 0.5%. This compares to a miscarriage rate of 1% for CVS. Many fetuses with severe genetic defects miscarry naturally during the first trimester.
• the trauma of difficult family-planning decisions. The threat posed to parental and family mental health from the trauma accompanying an abnormal test result can not be underestimated.

Normal results

Negative results from an amnio analysis indicate that everything about the fetus appears normal and the pregnancy can continue without undue concern. A negative result for Down syndrome means that it is 99% certain that the disease does not exist.

An overall "normal" result does not, however, guarantee that the pregnancy will come to term, or that the fetus does not suffer from some other defect. Laboratory tests are not 100% accurate at detecting targeted conditions, nor can every possible fetal condition be tested for.

Abnormal results

Positive results on an amnio analysis indicate the presence of the fetal defect being tested for, with an accuracy approaching 100%. Prospective parents are then faced with emotionally and ethically difficult choices regarding treatment options, the prospect of dealing with a severely affected newborn, and the option of elective abortion. At this point, the parents need expert medical advice and counseling.

Resources

PERIODICALS

Dreisbach, Shaun. "Amnio Alternative." Working Mother (March 1997): 11.

ORGANIZATIONS

American College of Obstetricians and Gynecologists. 409 12th St., S.W., P.O. Box 96920, Washington, DC 20090-6920. 〈http://www.acog.org〉.

OTHER

Holbrook Jr., Harold R. Stanford University School of MedicineWeb Home Page. February 2001. 〈http://www.stanford.edu/∼holbrook〉.

Amniocentesis

Definition

Amniocentesis is a procedure used to diagnose fetal defects in the early second trimester of pregnancy. A sample of the amniotic fluid, which surrounds a fetus in the womb, is collected through a pregnant woman's abdomen using a needle and syringe. Tests performed on fetal cells found in the sample can reveal the presence of many types of genetic disorders, thus allowing doctors and prospective parents to make important decisions about early treatment and intervention.

Purpose

Since the mid-1970s, amniocentesis has been used routinely to test for Down syndrome , by far the most common, nonhereditary, genetic birth defect, afflicting about one in every 1,000 babies. By 1997, approximately 800 different diagnostic tests were available, most of them for hereditary genetic disorders such as Tay-Sachs disease, sickle cell anemia, hemophilia, muscular dystrophy , and cystic fibrosis .

Amniocentesis, often called amnio, is recommended for women who will be older than 35 on their due-date. It is also recommended for women who have already borne children with birth defects, or when either of the parents has a family history of a birth defect for which a diagnostic test is available. Another reason for the procedure is to confirm indications of Down syndrome and certain other defects which may have shown up previously during routine maternal blood screening.

The risk of bearing a child with a nonhereditary genetic defect such as Down syndrome is directly related to a woman's age—the older the woman, the greater the risk. Thirty-five is the recommended age to begin amnio testing because that is the age at which the risk of carrying a fetus with such a defect roughly equals the risk of miscarriage caused by the procedure–about one in 200. At age 25, the risk of giving birth to a child with this type of defect is about one in 1,400; by age 45 it increases to about one in 20. Nearly half of all pregnant women over 35 in the United States undergo amniocentesis and many younger women also decide to have the procedure. Notably, some 75% of all Down syndrome infants born in the United States each year are to women younger than 35.

One of the most common reasons for performing amniocentesis is an abnormal alpha-fetoprotein (AFP) test. Alpha-fetoprotein is a protein produced by the fetus and present in the mother's blood. A simple blood screening, usually conducted around the 15th week of pregnancy, can determine the AFP levels in the mother's blood. Levels that are too high or too low may signal possible fetal defects. Because this test has a high false-positive rate, another test such as amnio is recommended whenever the AFP levels fall outside the normal range.

Amniocentesis is generally performed during the 16th week of pregnancy, with results usually available within three weeks. It is possible to perform an amnio as early as the 11th week, but this is not usually recommended because there appears to be an increased risk of miscarriage when done at this time. The advantage of early amnio and speedy results lies in the extra time for decision making if a problem is detected. Potential treatment of the fetus can begin earlier. Important, also, is the fact that elective abortions are safer and less controversial the earlier they are performed.

Precautions

As an invasive surgical procedure, amnio poses a real, although small, risk to the health of a fetus. Parents must weigh the potential value of the knowledge gained, or indeed the reassurance that all is well, against the small risk of damaging what is in all probability a normal fetus. The serious emotional and ethical dilemmas that adverse test results can bring must also be considered. The decision to undergo amnio is always a matter of personal choice.

Description

The word amniocentesis literally means "puncture of the amnion," the thin-walled sac of fluid in which a developing fetus is suspended during pregnancy. During the sampling procedure, the obstetrician inserts a very fine needle through the woman's abdomen into the uterus and amniotic sac and withdraws approximately one ounce of amniotic fluid for testing. The relatively painless procedure is performed on an outpatient basis, sometimes using local anesthesia.

The physician uses ultrasound images to guide needle placement and collect the sample, thereby minimizing the risk of fetal injury and the need for repeated needle insertions. Once the sample is collected, the woman can return home after a brief observation period. She may be instructed to rest for the first 24 hours and to avoid heavy lifting for two days.

The sample of amniotic fluid is sent to a laboratory where fetal cells contained in the fluid are isolated and grown in order to provide enough genetic material for testing. This takes about seven to 14 days. The material is then extracted and treated so that visual examination for defects can be made. For some disorders, like Tay-Sachs, the simple presence of a telltale chemical compound in the amniotic fluid is enough to confirm a diagnosis. Depending on the specific tests ordered, and the skill of the lab conducting them, all the results are available between one and four weeks after the sample is taken.

Cost of the procedure depends on the doctor, the lab, and the tests ordered. Most insurers provide coverage for women over 35, as a follow-up to positive maternal blood screening results, and when genetic disorders run in the family.

An alternative to amnio, now in general use, is chorionic villus sampling, or CVS, which can be performed as early as the eighth week of pregnancy. While this allows for the possibility of a first trimester abortion, if warranted, CVS is apparently also riskier and is more expensive. The most promising area of new research in prenatal testing involves expanding the scope and accuracy of maternal blood screening as this poses no risk to the fetus.

Preparation

It is important for a woman to fully understand the procedure and to feel confident in the obstetrician performing it. Evidence suggests that a physician's experience with the procedure reduces the chance of mishap. Almost all obstetricians are experienced in performing amniocentesis. The patient should feel free to ask questions and seek emotional support before, during and after the amnio is performed.

Aftercare

Necessary aftercare falls into two categories, physical and emotional.

PHYSICAL AFTERCARE During and immediately following the sampling procedure, a woman may experience dizziness, nausea , a rapid heartbeat, and cramping. Once past these immediate hurdles, the physician will send the woman home with instructions to rest and to report any complications requiring immediate treatment, including:

• Vaginal bleeding. The appearance of blood could signal a problem.
• Premature labor. Unusual abdominal pain and/or cramping may indicate the onset of premature labor. Mild cramping for the first day or two following the procedure is normal.
• Signs of infection. Leaking of amniotic fluid or unusual vaginal discharge, and fever could signal the onset of infection.

EMOTIONAL AFTERCARE Once the procedure has been safely completed, the anxiety of waiting for the test results can prove to be the worst part of the process. A woman should seek and receive emotional support from family and friends, as well as from her obstetrician and family doctor. Professional counseling may also prove necessary, particularly if a fetal defect is discovered.

Risks

Most of the risks and short-term side effects associated with amniocentesis relate to the sampling procedure and have been discussed above. A successful amnio sampling results in no long-term side effects. Risks include:

• Maternal/fetal hemorrhaging. While spotting in pregnancy is fairly common, bleeding following amnio should always be investigated.
• Infection. Infection, although rare, can occur after amniocentesis. An unchecked infection can lead to severe complications.
• Fetal injury. A very slight risk of injury to the fetus resulting from contact with the amnio needle does exist.
• Miscarriage. The rate of miscarriage occurring during standard, second trimester amnio appears to be approximately 0.5%. This compares to a miscarriage rate of 1% for CVS. Many fetuses with severe genetic defects miscarry naturally during the first trimester.
• The trauma of difficult family-planning decisions. The threat posed to parental and family mental health from the trauma accompanying an abnormal test result can not be underestimated.

Normal results

Negative results from an amnio analysis indicate that everything about the fetus appears normal and the pregnancy can continue without undue concern. A negative result for Down syndrome means that it is 99% certain that the disease does not exist.

KEY TERMS

Alpha fetoprotein (AFP) —A substance produced by a fetus' liver that can be found in the amniotic fluid and in the mother's blood. Abnormally high levels of this substance suggests there may be defects in the fetal neural tube, a structure that will include the brain and spinal cord when completely developed. AFP may also be found at elevated levels in the blood of adults with liver, testicular, and ovarian cancer.

Anencephaly —A genetic defect resulting in the partial to complete absence of the brain and malformation of the brainstem.

Chorionic villus sampling —A procedure used for prenatal diagnosis at 10–12 weeks gestation. Under ultrasound guidance a needle is inserted either through the mother's vagina or abdominal wall and a sample of the chorionic membrane. These cells are then tested for chromosome abnormalities or other genetic diseases.

Chromosome —A microscopic thread-like structure found within each cell of the human body and consisting of a complex of proteins and DNA. Humans have 46 chromosomes arranged into 23 pairs. Chromosomes contain the genetic information necessary to direct the development and functioning of all cells and systems in the body. They pass on hereditary traits from parents to child (like eye color) and determine whether the child will be male or female.

Down syndrome —A chromosomal disorder caused by an extra copy or a rearrangement of chromosome 21. Children with Down syndrome have varying degrees of mental retardation and may have heart defects.

Genetic —Refers to genes, the basic units of biological heredity, which are contained on the chromosomes.

Hereditary —Something which is inherited, that is passed down from parents to offspring. In biology and medicine, the word pertains to inherited genetic characteristics.

Maternal blood screening —Screening that is normally done early in pregnancy to test for a variety of conditions. Abnormal amounts of certain proteins in a pregnant woman's blood raise the probability of fetal defects. Amniocentesis is recommended if such a probability occurs.

Tay-Sachs disease —An inherited disease caused by a missing enzyme that is prevalent among the Ashkenazi Jewish population of the United States. Infants with the disease are unable to process a certain type of fat which accumulates in nerve and brain cells, causing mental and physical retardation, and, finally, death.

Ultrasonography —A medical test in which sound waves are directed against internal structures in the body. As sound waves bounce off the internal structure, they create an image on a video screen. Ultrasonography is often used to diagnose fetal abnormalities, gallstones, heart defects, and tumors. Also called ultrasound imaging.

An overall "normal" result does not, however, guarantee that the pregnancy will come to term, or that the fetus does not suffer from some other defect. Laboratory tests are not 100% accurate at detecting targeted conditions, nor can every possible fetal condition be tested for.

Abnormal results

Positive results on an amnio analysis indicate the presence of the fetal defect being tested for, with an accuracy approaching 100%. Prospective parents are then faced with emotionally and ethically difficult choices regarding treatment options, the prospect of dealing with a severely affected newborn, and the option of elective abortion. At this point, the parents need expert medical advice and counseling.

Parental concerns

There is a risk of miscarrage with this procedure.

When to call a doctor

If there is excess bleeding, a doctor should be contacted.

Resources

BOOKS

Hassold, Terry and Schwartz, Stuart. "Chromosome Disorders." In Harrison's Principles of Internal Medicine, ed. Eugene Braunwald, et al. Philadelphia: McGraw-Hill, 2001.

Miesfeldt, Susan and Jameson, J. Larry. "Screening, Counseling, and Prevention of Genetic Disorders." In Harrison's Principles of Internal Medicine, ed. Eugene Braunwald, et al. Philadelphia: McGraw-Hill, 2001.

Wallach, Jacques. Interpretation of Diagnostic Tests, 7th ed. hiladelphia, PA: Lippincott Williams & Wilkens, 2000.

ORGANIZATIONS

American College of Obstetricians and Gynecologists. 409 12th St., S.W., P.O. Box 96920, Washington, DC 20090-6920. <http://www.acog.org>.

ORGANIZATIONS

National Institutes of Health. <http://www.nlm.nih.gov/medlineplus/encyclopedia.html>.

Mark A. Best