prenatal diagnosis in the 1990s

Prenatal Diagnosis in the 1990s ~ ~ ~~

Lynette Wright, RN, MN

Prenatal diagnosis has become widely available and detects an increasing variety of birth defects and potentially harmful medical conditions. Many of the studies are complex and must be performed within a specific time period. Most prenatal diagnostic sampling techniques have some degree of risk for the mother or the fetus, and all produce at least transient anxiety. Nurses are involved in identifying families at risk, preparing women for the procedures, providing support, and counseling patients after the results are known; because of this role, nurses need updated information. This review describes current methods for early identification of a potential problem, discusses a variety of prenatal diagnostic procedures, reviews the most common types of laboratory studies, and introduces future trends in the field of prenatal diagnosis.

n very pregnancy carries with it a 3-5% chance of significant birth defects. In addition, many families

have specific risk factors that increase their chances for delivering a child with physical abnormalities or mental retardation. These include maternal diseases; drug or chemical exposures; genetic disorders identified in the family history; or maternal age, which increases the risk for a chromosomal abnormality. An increasing number of families are seeking prenatal tests to determine the health of the developing fetus.

Prenatal screening and diagnosis allows couples at risk for a specific defect to test for the presence or absence of an abnormality, provides a range of informed choices for parents, provides reassurance, and reduces anxiety for couples at risk. .Reassurance is an important reason for prenatal diagnosis. In more than 98% of cases, the prenatal diagnostic tests will have normal results for the condition in question. Only 2% will have findings that confirm serious abnormalities in the fetus. Although in the minority, these couples will have to process some difficult options and make significant decisions in a short period of time (Thompson, McInnes, & Huntington, 1991).

It is important that a nurse caring for families during a pregnancy be able to identify potential problems for which prenatal diagnosis is available, understand current prenatal diagnostic techniques, and be aware of the trends for the future. The nurse also must appreciate the psychological dynamics that are present when a couple is considering prenatal testing.

Identifying a Potential Problem

When we think of prenatal studies, the nursing literature tends to emphasize expensive esoteric tests. However, equally important are routine prenatal screening procedures regularly provided by obstetric practices. The three most common obstetric screening procedures for identifying genetic problems are family history, alpha-fetoprotein (AFP) and multiple marker screening, and ultrasound evaluation.

Family History Obtaining a family history is considered routine, yet it is the single most important tool for assessing genetic risk. As the genes are mapped to specific human chromosomes and molecular testing becomes more widely available, family history assessment will assume a critical role in determining which molecular tests are appropriate. Currently family history is the primary means for identifying pregnant women who need genetic counseling and perhaps special prenatal studies for potential chromosome disorders, single gene conditions, or multifactorially caused birth defects. A good family history identifies the following problems:

1. Advanced maternal age. In the United States a growing number of women are pursuing full-time ca- reers. Economic and other sociologic changes also have dramatically increased the age at which many women are beginning their families. Nurses should recognize that although maternal age is the greatest indicator for amniocentesis, older mothers are only at increased risk for chromosome misdivision. They do not have an increased

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Maternal Age

risk for birth defects or mental retardation caused by other genetic mechanisms.

Advanced maternal age is defined as women who will be 35 years or older at delivery. These women have an increased risk for chromosomal problems in the fetus because of misdivision of the ovum during meiosis. The likelihood that an ovum will misdivide begins to increase at approximately 28 years. This risk increases slowly until the mid-30s and then increases at a faster rate (see Fig. 1). Age 35 was chosen as a cut-off point for age-related amniocentesis only because early studies in the 1970s showed that by age 35, the risk for a chromosome problem exceeded the risk of the amniocentesis procedure. Offering amniocentesis to women 35 years of age and older has been the standard of care for almost 20 years. However, since that time, the introduction of ultrasound, disposable amniocentesis trays, and increased experience have reduced the risk of the amniocentesis procedure, and many experts think women in their early 30s could be included. However, other less expensive and less invasive screening tests have been developed that may meet the needs of younger women, including women between 30 and 34 years of age.

2. A previous child with any chromosome abnormality. Couples who have had a child with Down syndrome, trisomy 18, trisomy 13, or Turner syndrome often seek prenatal diagnosis for subsequent pregnancies. Such parents usually have heightened anxiety, even if their statis- tical risk for recurrence is low. In addition, data based on studies of parents of trisomy 21 Down syndrome children indicate a recurrence risk of 1 in 100, even when the mother is younger than 30 years. This is eight times higher than the age-related population risk and may be caused by gonadal mosaicism in some of these parents. Because such parents would have normal blood, chromosomes and mosaicism in ova or sperm usually are not detectable, prenatal diagnosis is an important option for any

couple with a previous pregnancy that was chromosom- ally abnormal (Thompson et al., 1991).

3 . A known chromosome translocation or other structural chromosome rearrangement in either parent. Func- tionally normal parents also can carry chromosomes that have broken and reattached. Although parents with a balanced translocation will be mentally and physically normal, a problem occurs when their chromosomes separate at meiosis. Depending on how the individual chromosomes line up and divide, resultant ova and sperm can be normal, balanced (rearranged but with the correct amount of genetic material), or may be left with too much or too little genetic material. If the gametes have an abnormal amount of genetic material, the most frequent outcome is spontaneous abortion. In other cases, these rearrangements also result in live-born children with chromosome abnormalities. Thus, when a family history reveals multiple miscarriages, stillbirths, or children with abnormalities and a cause cannot be precisely determined, a referral for genetic counseling and parental chromosome studies should be made.

When a couple has had multiple miscarriages they may have had parental chromosome studies or fetal chromosome studies in the past. In many cases, families had testing during a time of crisis or acute grief and may not remember the reason testing was done or the results. They may feel guilty because they “carry” an abnormal chromosome arrangement. The nurse should use some open-ended questions to determine their understanding and their feelings. Clarification or referral for genetic counseling may be needed.

4 . Family history of a known genetic syndrome or any disorder “repeating” in a family. Many families will be aware of a family history of a particular genetic disorder. In some dominant conditions, many affected family members will be identified, whereas other dominant conditions, such as Marfan syndrome, deafness, and neu- rofibromatosis, may be variable and may not be recog- nized as “genetic” by the family. Dominant conditions also can be new mutations and may look like an isolated case until that person reproduces. Achondroplastic dwarfism is a good example of a frequent new mutation.

In autosomal recessive conditions such as phenylke- tonuria (PKU) and sickle cell anemia there may be no previous history of an affected person. Because two genes must be transmitted, and one from each side of the family must be present to produce medical problems, it is likely that the disorder will express itself in onlyone individual or within a single sibship. A potential risk for certain recessive disorders can be determined by identifying the ethnicity of a couple. For example, Tay Sachs and Gaucher’s diseases are more common in persons of Eu- ropean Jewish descent, whereas cystic fibrosis and alpha one antitrypsin disorder are more common in persons of northern European ancestry.

X-linked recessive disorders such as Duchenne muscular dystrophy or hemophilia may be well delineated within the family or may be noted by pedigree analysis. Because there are several X-linked disorders that can be

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diagnosed prenatally, it is critical that the nurse recognize early i n pregnancy any family history with a pattern o f affected males. Such Pamilies should he immediately referred for genetic counseling a n d medical evaluation.

When there is a family history o fa genetic syndrome, the nurse often may encounter siblings or other relatives who are concerned about their risks and are seeking prenatal diagnosis. With the emergence o f DNA techniques, an increasing number o f single gene disorclers can be di- agiiosed during fetal life. To make an appropriate referral, it is important that the nurse delineate the type of disorder, determine which family members are affected, ask ;ibout clinical variability among rekitives, and clefine any known pattern o f inheritance.

5. History of birth defects in close family members. Defining “birth defects” cmi be difficult. Many different modes o f inheritance can produce birth defects, whereas other defects can be caused by teratogenic agents or can be sporadic developmental errors. Many isolated birth defects, such as spina bifida, cleft lip and palate, and \’en- tricular septa1 defects, are :I combin:rtion o f predisposing genetic factors and intrauterine environmental influ- ences. These multifactorially inherited conditions are identified primarily by ultrasound, but emerging DNA techniques are expanding our ability t o provide precise diagnoses.

Routine Maternal Serum AFP and Multiple Marker Screening

Maternal serum screening began as screening for spina hifida. Alpha-fetoprotein is made by the fetal yolk sac and fetal liver and generally is present throughout the fetus. I t is processed and excreted i n t o the amniotic fluid, swal- lowed, and reprocessccl by the fetus. A small amount crosses the placenta and can be measured in maternal blood. If :I major o t x n defect such as snina bifida or om- phalocele is present, a greater amount is excreted into the amniotic sac, and a proportionally higher amount crosses the placenta. Thus, the AFP level in maternal serum is elevated.

I t was noted that when the AFP value was low, a higher-than~expected number o f infants with Down syndrome were identified. However, AFP alone identifies only approximately 20% o f Down syndrome fetuses and will miss approximately 80% o f cases.

Because AFP alone is inadequate t o detect Down syndrome, research was begun o n other annolytes present in early fetal development. l’wo of these. human chorionic gonadotropin (hCG) a n d tinconjugated estriol (uE3) also are altered when Down syndrome was present. By combining analysis o f maternal age, AFP, hCG, and uE3 values and adjusting for variations in weight, race, and diabetic status, a multiple marker screening test has evolved that can identify women who need additional evaluation for carrying a fetus with 1)own syndrome. Multiple marker screening identifies apprc;ximately 70% of Down syndrome cases, and new refinements have

made it possible to detect trisomy 18 on the same blood specimen and t o indicate risks for trisomies in twin pregnancies (Canick & Knight, 1991).

When there is a positive AFP or multiple marker screening test that cannot be resolved by ultrasound &at- ing, identification of multiple gestation, or information regarding maternal complications, an amniocentesis should be considered. Amniocentesis and high-level ultrasound are the back-up diagnostic tests for AFP and multiple marker screening procedures.

Ultrasound Evaluation Perhaps the most widely used prenatal screening technique for assessment of fetal well being is ultrasonogra- phy. A level I o r dating ultrasound confirms gestational age, identifies the location of the placenta, and checks for major fetal structures. Large birth defects such as anen- cephaly or severe dwarfism can and should be detected by a level I ultrasound. Level I ultrasounds are rarely diagnostic when used alone. If there is a family history of birth defects, any indication of growth problems or an increased risk for birth defects in the existing pregnancy, a level 11 ultrasound and evaluation by an obstetric or perinatal specialist woulct be needed.

A prenatal level I 1 ultrasound is a systemic scan of the body systems, including the cranium, thorax, heart, spinal column, limbs and digits, performed by an opera- tor experienced in detecting fetal anomalies. Informa- tion regafding intrauterine growth and proportions for the particular gestational age are included. A single level I1 ultrasound often is not diagnostic without other con- firmation, but it can identify higli-risk pregnancies that are in need of special diagnostic tests, serial ultrasound studies, o r alterations in pregnancy management (see Ta- ble 1). The most distinguishing feature between a level I and a level 11 ~iltrasound is the experience and the training of the provider who is interpreting the scan.

Prenatal Diagnostic Procedures Prenatal diagnosis began in the later 1960s when techniques were developed to isolate and culture fetal cells

Table 1. Ultrasound Findings Commonly Associated with Chromosome Abnormalities or Genetic Syndromes

Shortened femur length Thickened nuchal fold Cystic h ygroma

Pol ycyst i c kidneys Skeletal dysplasias Neural tube defects Congenital heart defects Cleft lip and palate Intrauterine Growth Retardation

Pol ydactyl y

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from amniotic fluid. At that time the safety and efficacy of amniocentesis had not been established. The procedure could not be performed until 16 weeks’ gestation because 25-30 mL of fluid was required, and results were not available for about 4 weeks. Since that time improvements i n laboratory methods have enabled a coniprehen- sive analysis with a smaller sample and with results available in less than 2 weeks.

During the past 20 years, a variety of techniques have developed to identify anomalies and assess for fetal well- being. In the past, most of these techniques involved a degree o f risk and were not available until the 2nd trimester. In recent years, many refinements have been developed in response to public demand for earlier and faster results and for less-invasive techniques. A variety of procedures for obtaining fetal cells have emerged, including chorionic villi sampling, early amniocentesis, traditional amniocentesis, and cordocentesis. Each o f these procedures has specific benefits and limitations, which are detailed in this article.

Traditional Amniocentesis

Traditional amniocentesis is the safest and most widely used prenatal diagnostic technique and has been used in the United States for more than 20 years. Traditional amniocentesis is performed between 15 and 18 weeks’ gestation.,Ultrasound is completed before the amniocentesis procedure to identify the location of the fetus, the placenta, and the best pocket of fluid in which to place the amniocentesis needle. The procedure usually is performed by inserting a needle transabdominally and with- drawing about 20 mL of amniotic fluid from the sac sur- rounding the fetus.

The entire amniocentesis process takes about 20 minutes, including completing the ultrasound, applying sterile drapes and antiseptic solution t o the abciomen to prevent infection, and obtaining the fluid sample. With- drawing the amniotic fluid takes about the same amount of time it takes to draw a routine blood saniple and has about the same level of discomfort. Some women may feel a cramp during the procedure, but more often women describe a “stick” on the surface and pressure or a “push-pull” feeling when the fluid sample is withdrawn (see Fig. 2).

The risk for any type of complication, including vagi- nal spotting, leakage of amniotic fluid, significant cranip- ing, infection or miscarriage within 48 hours of the procedure is less than 0.5% (0.2-0.4%) when the procedure is done by experienced clinicians (D’Alton B DeCher- ney, 1993). However, because this is an outpatient procedure, it is important that the nurse caution the patient to call immediately if bleeding, leakage of fluid, fever, significant pain, or cramping occur. Certain activities should be limited for 48 hours to allow the puncture site to repair and the fluid to be replaced. These precautions are listed in Table 2 .

Early Amniocentesis Because of advances in laboratory techniques and im- proveci ultrasound guidance, amniocentesis now can be eflectively performed at 12-14 weeks’ gestation. Early amniocentesis allows the patient another choice for prenatal diagnosis and decreases the period of patient anxiety. Chromosome results can be obtained within 10-14 days with 99.9% accuracy. The risk for culture failure is similar to that associated with traditional amniocentesis when the procedure is performed by a11 experienced cli- nician, although earlier national studies showed a 0.2- 0.3% national culture failure rrtte (D’Alton B DeCherney, 1993).

Early amniocentesis is performed in the same man- ner as traditional amniocentesis. Approximately 1 mL of amniotic fluid per gestational week is obtained. Because of the smaller volume of fluid ]>resent at 12-14 weeks’ gestation, there is a slightly increased risk for obtaining an inadequate sample. However, laboratories that use a coverslip method should be able to perform all necessary chromosome studies with 12-15 mL of amniotic fluid. The risk of complications such as miscarriage after early amniocentesis is approximately I%, which is slightly greater than the risk of traditional amniocentesis. As with

Table 2. Precautions after Amniocentesis or CVS ~ ~~

1 . Trtke the rest of the day off, if possible. 2. Rest (get off y o u r feet) for a couple o f hours (not

necessarily immediately after the procedure) 3. Avoid heavy lifting (more than 20 pounds) for several

4 . Avoid bending at the waist 5 . Avoid strenuous exercise o r activity 6 . Report any fever, fluid leakage, cramping o r spotting

days

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all other prenatal diagnostic procedures, earlyamniocen- tesis risks decrease as the technical experience of the physician increases.

Early amniocentesis may be contraindicated because of anatomical differences. If there is a complete anterior placenta, a large anterior fibroid, or a severely retroverted uterus, appropriate needle placement may not be possible until the uterus is larger. Women weighing more than 180 pounds may be too large in proportion to the size of the uterus. In twin pregnancies, each amniotic sac would be too small and have too little fluid at 12-14 weeks’ gestation to obtain a safe sample.

Initially, AFP testing on amniotic fluid earlier than 14 weeks’ gestation was not possible. Normative data have been compiled for these gestational weeks, but detection rates are lower than at 16-18 weeks’ gestation. Acetylcho- linesterase (ACHE) can be used with amniotic fluid AFP from 11 to 15 weeks’ gestation to improve detection rates. Positive results o n AFP and ACHE testing are predictive, for spina bifida, but a negative result may be less informa- tive. When both tests are used at 13-15 weeks’ gestation, the detection rate is 90%. Both tests also should be used for 11-12-week amniocentesis. ACHE is not an effective measure at less than 10 weeks because circulating ACHE from the period before the closure of the neural tube re- mains (Wald, Cuckle, & Nanchahal, 1989).

Chorionic Villus Sampling

Chorionic villus sampling (CVS) is a prenatal diagnostic technique performed in the 1st trimester of pregnancy. This diagnostic technique allows the patient to make early decisions concerning the pregnancy and helps to lessen patient anxiety.

CVS can be performed between 9 and 12 weeks’ gestation and is possible because the amnion, the chorion, and the three-layer embryo are all fetal in origin and thus have the same chromosome and DNA constitution. The chorion develops from trophoblasts, which by the e n d of the 8th week cover the entire surface of the chorionic sac. The part of the villi that attaches to the wall of the uterus will develop into the placenta, and after about 11 weeks, the remainder of the villi will begin to degenerate and the chorion will become smooth. Because the villi are fetal in origin, one has a window of time in which a sample of villi can be safely removed and tested (see Fig. 3).

I t is safest to perform CVS between 10 and 11.5 weeks’ gestation; a 9- or 8-week CVS is possible, but there is a higher risk for miscarriage and a possible increased risk for some types of birth defects. During the 9-12- week period, chorionic villi can be aspirated transvagi- nally or transabdominally. When CVS is performed by experienced clinicians, the fetal loss rate is 2.3% for trans- abdominal CVS and 2.5% for the transvaginal procedure. Two randomized trials demonstrated that women as- signed to a 1st trimester CVS group had a 1.7-4.6% greater chance for an unsuccessful outcome than did the women in the amniocentesis group. Approximately 5% of patients having CVS may deliver 1-2 weeks early (Cana-

l

I Figure 3. Vuginul choriariic rillus’ sampling procedure.

dian Collaborative, 1989). Other complications include infection, spotting, bleeding, cramping, and leakage of amniotic fluid.

Recently, public media has focused o n the possibility of limb reduction defects after the CVS procedure. A Brit- ish study in 1991 reported 5 limb defects and oroman- dibular hypoplasia in several infants whose mothers had undergone CVS procedures between 56 and 66 days after their last m‘enstrual period. A United States study by Bur- ton, Schulz, and Burd (1992) also notes transverse finger and toe defects after chorionic villus sampling. Other studies, including a large study by Mahoney showed n o increased incidence. Timing of the procedure and physician experience may be critical factors. Although the proposed mechanism is a vascular insult leading to fetal hypoperfusion, a causal relationship between CVS and limb defects has not been established. Clinical research is ongoing (Burton et al., 1992; D’Alton & DeCherney, 1993).

Chromosome analysis of chorionic villi should be completed within 5-14 days, depending on the culture procedure used. Early literature discussed a 1-3-day result. These were direct preparations in which studies were performed immediately after the procedure or after an overnight incubation. These results are less accurate and show a higher rate of false mosaicism. For these reasons, any direct preparation result should be considered preliminary. Many laboratories have stopped reporting direct preparations. Final CVS results should be from a cultured cell analysis.

The overall accuracy rate of CVS chromosome analysis is reported to be approximately 94-96% because of an increased risk for maternal cell contamination and a phenomena called placental mosaicism. True mosaicism is the presence of both normal and abnormal cell lines in a fetus or individual because of nondisjunction (cell misdivision) that occurred at some point after fertiliza- tion. Placental mosaicism occurs after the development

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of the extra embryonic tissues and is not associated with fetal abnormalities. About 1% of CVS studies will have results that indicate possible mosaicism. In these cases the chromosome results may be inconclusive and require amniocentesis at 14-16 weeks for definitive results (D'AI- ton bi DeCherney, 1993).

There are limits to the diagnostic tests that can be performed using CVS. I t is diagnostic for chromosome abnormalities and a few other genetic disorders that can be identified by DNA analysis. Alpha-fetoprotein testing for neural tube defects is not available with CVS because AFP must be analyzed directly from amniotic fluid. Thus, a woman having CVS would require maternal serum AFP testing and ultrasound evaluation at 15-18 weeks' gestation. An amniocentesis would be needed for additional evaluation if maternal serum AFP was elevated.

Although CVS carries a higher risk for complications than do early or traditional amniocentesis, it has developed in response t o public demand for earlier prenatal diagnosis. As we move into the future, the demand for 1st trimester studies will grow, as will the demand for less- invasive procedures.

C o rdocen tesis The cordocentesis procedure uses ultrasound guidance to obtain a sample of fetal blood directly from the umbil- ical cord. This also is known as PUBS (per cutaneous um- bilical blood sampling) and is the most invasive of the prenatal procedures.

Cordocentesis is used when it is necessary to have actual fetal blood for analysis, such as in certain enzyme tests, or when faster results are needed because of late gestational age or known ultrasound abnormalities. Cor- docentesis is required less frequently because an increasing number of diagnostic tests can be performed using DNA from cultured amniotic cells, and in situ hybridization techniques can provide rapid chromosome analysis for some chromosome pairs. For example, prenatal diagnosis for sickle cell anemia required PUBS in the past. Now a direct DNA test is available, and sickle cell anemia can be diagnosed with fetal cells obtained from CVS or amniotic fluid. The nurse should check with an established genetic center or laboratory when a cordocentesis is anticipated to see if newer diagnostic techniques have made a less-invasive procedure possible.

Types of Laboratory Studies Chromosome, DNA, and in situ hybridization studies are performed on material taken from the nucleus of fetal cells, which can be obtained by any of the CVS or amniocentesis procedures. Biochemical studies may require the amniotic fluid for analysis because such studies often measure the quantity of an enzyme or annolyte in the fluid portion.

Chromosome Analysis Chromosome analysis can be performed by obtaining any tissue that contains cells capable of dividing. Usually the

Figure 4 Chromosomal kuiyo(ipe

biological sample is blood or amniotic fluid fetal cells. These cells are grown in culture until there is sufficient quantity for analysis, usually 2-3 days for blood and 4-6 days for an amniotic fluid sample.

When cells are not dividing, the chromosome material is unwound, much like a plate of spaghetti, and individual chromosomes cannot be counted. At that time, the metabolic work of the cell is accomplished. As the cell prepares to divide, the individual chromosomes become compact, like thick macaroni, and can be individually counted. This stage is known as metaphase.

A cell population with multiple colonies and numer- ous cells (usually a minimum of 15 colonies with 20 or more cells in each colony) must be established. Because individual cells are dividing at different times, it is critical to have a large enough population of cells to be able to identify enough metaphases so that chromosomes can be counted and analyzed.

Once an adequate cell population is established, a chemical is applied that stops all cell growth and divi- sion. Other chemicals are added to spread the chromosomes and band them so that each chromosome can be individually identified. Counted metaphases are photo- graphed, cut out, and arranged in a pattern that is called a karyotype (see Fig. 4). Computers are used in this process, which until recently was done hand.

Karyotypes are analyzed and reviewed by experienced technicians and laboratory directors before a report is issued. The entire process should take about 7-10 days to complete.

Biochemical Analysis More than 100 metabolic disorders, such as Tay Sachs disease, galactosemia, and methylmalonic acidemia, can be diagnosed prenatally by analysis of CVS tissue, cultured amniocytes, or assay of a substance in the amniotic fluid. Most of these disorders are rare and usually recessive,

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which means that although they occur infrequently in the general population, they have a high recurrence risk (usually 25%) within sibships.

It may not be possible to test for some biochemical disorders because the substance to be measured, such as a liver enzyme, may not be expressed in amniocytes or amniotic fluid. These must be addressed by the newly de - veloping array of DNA tests. In other instances, biochemical testing has distinct advantages. Biochemical analysis may be advantageous when there are many mutant alleles or frequent new mutations that affect the function of the protein. The biochemical test measures whether adequate enzyme or other product is being made, regardless of the genotype (Thompson et al., 1991).

Because most biochemical conditions are rare, the experience of the laboratory performing the test is important. N o laboratory or genetic center will perform more than a few of these tests. It often is in the patient’s best interest to obtain the sample locally, culture cells, and send a sample t o a center specializing in one area of biochemical research. In many instances, the referral laboratory requires cultured cells and will not accept CVS tissue or amniotic fluid samples directly. Thus, it is important to refer to a genetic laboratory with expertise in biochemical testing and collaborative relationships with a variety of specialty and research centers. The nurse should make sure that n o prenatal sample is drawn until the protocol for sample collection (sample type, sample size, and handling of specimen) and analysis is known.

DNA Analysis A variety of disorders that were not previously detectable can be diagnosed using DNA analysis. This trend will continue and will be extended t o include behavioral and psychiatric conditions; common diseases, such as cancer, stroke, anti diabetes; and late-onset disorders, such as Alzheimer’s disease. DNA analysis will become a major diagnostic modality within the next decade. Thus, the nurse must understand the concepts relating to DNA analysis beyond the conditions that now can be prenatally diagnosed by this method.

Understanding DNA analysis requires a brief biological review. Each nucleated cell in the entire body contains almost all o f the genetic information for an entire individual. The genomic information is housed on linear strands of DNA, which are made up of individual base pairs arranged in a precise order. Groups of base pairs make up to 100,000 or more individual genes that code for specific proteins in humans. To encode an enormous amount of information into each individual cell, the DNA must be twisted, coiled, and. supercoiled in a miracle of micropackaging. Precise packaging o f DNA also provides control over which genes are expressed at a given time.

Biological health depends on species variability, which in humans is encouraged by several natural mechanisms. These include recombination and assortment during meiosis and the introduction of permanent changes through mutation. Some mutations significantly alter the structure and function o f a protein and cause a

genetic disorder. Other mutations are not harmful. In fact, the presence of several genetically different healthy forms of a gene, called polymorphism, is evidence for species adaptation. Such polymorphisms are common and allow the laboratory to distinguish different inherited forms of a gene within family groups.

The two broad categories of DNA analysis are direct and indirect detection. Each category uses multiple laboratory methods, such as Southern blotting o r polymerase chain reaction techniques, but the type of analysis depends on how much is known about a given gene and the particular mutation in question. For example, suppose a patient is a known carrier for Duchenne muscular dystrophy, and the entire family has had DNA testing. If the mutation that caused the muscular dystrophy is a deletion, prenatal diagnosis is possible by a direct method. If the Duchenne muscular dystrophy is caused by a gene dupli- cation, a direct study is not possible, but prenatal diagnosis is available by family studies and linkage analysis.

Direct detection is possible when the gene has been located on a particular chromosome and the type and ex- act location of alteration that has occurred is known. Di- rect testing is desirable because direct DNA testing can be accomplished using blood or tissue from a single individual. Direct detection does not require blood or tissue samples from affected relatives or even a known family history of the disease. Direct detection is highly accurate but tests only for precise mutations. For example, it is possible to use direct DNA methods to test for carriers or affected persons with cystic fibrosis (CF). Such methods are available for between 16 and 20 CF mutations, including those that most commonly cause severe CF. However, since 1989, more than 250 CF mutations have been identified, including many that are quite rare. I t would be impractical to develop a battery of direct tests that would eliminate the risk for transmitting a CF gene.

Indirect testing or linkage analysis is required when the gene location or the precise alteration of a particular mutation is not known. In such situations, the gene can be traced through family studies. This is possible because genes that are close together usually are inherited together. Most families have polymorphisms (normal variations in their DNA) that are detectable. If polymorphisms can be detected that are close to the gene in question, the identifiable polymorphisms can be used to track the inheritance of the normal gene variations within a given family and predict who might have inherited the problem gene. Thus, linkage analysis is specific to each individual family and requires testing of several family members, usually including at least one person who is affected with the disease in question.

The accuracy of linkage testing depends on how close the markers are to the disease-causing mutation. The best polymorphisms would be those that closely flank the gene or intragenic markers. Results are expressed as probabilities.

Because of the preciseness of DNA analysis, i t is useful in all of the following situations:

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1. carrier detection; 2 . presymptomatic diagnosis; 3. prenatal diagnosis; 4. paternity and forensic identity testing; 5 . donor recipient matching; and 6 . evaluation for predisposition to late onset diseases,

i.e., cancers, stroke, diabetes.

However, testing is complex and variable, depending o n the precise family situation. Genetic counseling must be individualized for each family. In almost every case there is a possibility for an uninformative or incom- plete result. In other cases, a positive result may have life- long medical or psychological implications. Thus, when a DNA analysis is being considered, the family should be referred for an in-depth family history and counseling by a trained genetic counselor or medical geneticist.

Fluorescent In Situ Hybridization

It is possible to develop highly specific DNA sequences (probes) that will bind to their complimentary DNA strands in blood or tissue samples. A fluorescent label can be attached that enables the laboratory to quickly scan for the probe. Such advances in molecular cytogenetics have made it possible to identify a variety of chromosome abnormalities within 48 hours by in situ hybridization. Probes also are being developed to detect gene sequences associated with Mendelian disorders. The rapid chromosome detection technique is commercially available to detect numerical chromosome abnormalities in amniotic fluid cells for chromosomes 13, 18, 21, X , and Y. In the laboratory, fluorescent-labeled chromosome- specific DNA probes are used that bind (hybridize) to similar pieces of DNA in interphase (nondividing) cell nuclei. Thus, cells do not have to be cultured for 3-6 days before analysis can begin. The original sample is hybrid- ized for 12-24 hours and analyzed for fluorescent signals (see Fig. 5). The signals can be counted under a fluorescent microscope to determine if a normal chromosome

count or chromosome aneuploidy is present for a given chromosome pair (see Fig. 6) (Ward et al., 1993).

This test typically is requested when rapid results will improve patient care. Indications include fetal anomalies identified on ultrasound, maternal serum screening results that indicate an increased risk for Down syndrome or other chromosome abnormalities, a family history of previous trisoniies, or extreme parental anxiety. Because only five chromosome pairs are tested by the rapid method, in situ hybridization for numerical chromosome abnormalities is always offered in conjunction with a complete chromosome analysis.

There are several benefits to in situ hybridization studies. Only 5 m L o f additional amniotic fluid is required to perform i n situ hybridization studies. Informa- tive results are highly accurate. When the results are normal, anxious patients can be reassured within 48 hours. When there is an abnormal result, medical management plans, genetic counseling, and support services can be put in place. Staff training can take place in anticipation of a child with an ahnormality.

There also are limitations t o in situ hybridization technology. If there is blood in the sample, maternal cell contamination is possible, and the sample cannot he run. I f there is late gestation, debris in the amniotic fluid may have fluorescent properties, making analysis impossible. If too little fluid is submitted, the laboratory must opt for a complete chromosome analysis without the rapid in situ test. For these and other technical reasons, 8-10 of 100 women will receive an uninformative result and will have to wait for the results of their routine chromosome analysis. Thus, :my family opting for in s i tu hybridization studies should have adequate counseling regarding the benefits and limitations of this new technology.

Research is ongoing regarding other uses for in situ hybridization. One excellent use is to detect tiny dele- tions and point mutations that have clinical significance but are too small to be seen by even the most sophisti-

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cated chromosome analysis. When probes are developed f o r a precise sequence of DNA, the in situ method can be used with any sample to identify the presence o r absence of that particular sequence. In situ hybridization techniques also are used to detect structural and numerical abnormalities in leukemia, lymphoma, and solid tumor cells and is particularly useful when only a few cancer cells are present. Other potential uses include detection of particular genetic changes in an egg or sperm or in the very early embryo. As this technology progresses it will vastly expand o u r ability to provide prenatal diagnostic information (Gray et al., 1992).

Future Trends

The ability to detect and perhaps treat an expanding number of genetic conditions during the prenatal period is increasingly possible because of the worldwide collabo- ration known as the Human Genome Project. This project is attempting to map all human genes to precise locations on chromosomes, to develop methods for sequencing these genes, and to study the social ramifications of the emerging genetic technology.

Future improvements in prenatal diagnosis will continue to be driven by the desire for earlier and more precise diagnoses and for less-invasive methods to accom- plish these goals. Advances in molecular genetic technology have made two new methods, fetal cell diagnosis from maternal blood and preimplantation genetic diagnosis. feasible in the near future.

Fetal Cell Diagnosis Fetal cells cross the placenta and are circulating in maternal blood. Thus, it should be possible to collect some of these cells and analyze them without performing an invasive procedure that might place the pregnancy at risk. The research challenges include determining the most appropriate type of fetal cell, determining if these cells remain from previous pregnancies, developing methods to concentrate the fetal cells for analysis, and refining in situ hybridization and other molecular techniques for a rapid and cost-effective analysis. I t has been shown that trisomy 21 is detectable by this method. Research is ongoing to develop the technology and study the effective- ness of this method of prenatal diagnosis (Bianchi et al., 1993; Bianchi et al., 1992).

Preconception or Preimplantation Genetic Diagnosis High-risk families often have extreme anxiety, and wait- ing for a diagnostic procedure until late in the 1st trimester is difficult. Other families. have religious or cultural beliefs that would not allow a medical abortion for any fetal anomaly. For these families, it will be possible to use the molecular genetic techniques previously discussed to identify the presence or absence of many single gene and chromosome disorders in egg or sperm before conception or in the early embryo before organogenesis. The principle behind this technique is that every cell contains the entire genome for an individual. Thus, it

should be possible to take a single cell (or very few cells) and apply genetic probe technology to identify a particular gene. This technology would look for a known mutation for a particular single gene disorder. In situ hybridization techniques also might be used to detect a numerical chromosome abnormality.

It is possible in a few specialty research centers to obtain an ovum and remove the polar body for study. When gametes are formed, half of the paired genetic material goes to the egg, and the other half is in the polar body. If the polar body contains the mutation that the family wants to avoid, the egg should have the normal gene. If the polar body is normal, the problem gene should be in the egg. Normal eggs would be fertilized using in vitro techniques and implanted in the uterus, thereby avoiding the disorder in question.

Conclusion

Prenatal diagnosis is becoming more complex. It is important that nurses be knowledgeable about risk factors and the available testing procedures. However, it will be impossible for a general practice nurse, obstetrical nurse practitioner, or nurse midwife to keep up with all of the emerging technology. Every nurse should develop a genetic support system that includes laboratories with spe- cialized expertise in genetic testing, genetic counseling resources, and a network of support groups for families with fetal losses or genetic abnormalities. This support network wih allow the nurse to provide the most appropriate risk assessment, information, support services, and coordination of care.

Prenatal applications of genetic technology will continue to expand. However, technology should not drive the decision-making process. Some families will not want testing. Other families will want a diagnostic procedure knowing that if the results are abnormal they will continue the pregnancy and plan for the care of a child with special needs. Still other families will desire an early diagnosis in order to end an abnormal pregnancy. Other families will have many conflicts and will have difficulty making a decision. Thus, nurses should be nonjudg- mental in their approach. Whether counseling is done by a nurse or by a genetic specialist, the process should include nondirective strategies that allow families to ex- plore all of their options and make choices that fit their family needs and goals. All families deserve a caring ad- vocate who can support them during this decision-making process. The nurse often is in the best position to provide this support (Chesheir & Cefalo, 1992).

References

Bianchi, D. W., Mahr, A, , Zickwolf, C. K., Houseal, T. W., Flint, A. F., & Klinger, K. W. (1992). Detection of fetal cells with 47 XY + 21 karyotype in maternal peripheral blood. Hu- man Genetics, 90,368-370.

Bianchi, D. W., Zickwolf, G. K., Yih, M. C., Flint, A. F., Geifman, 0. H., Erikson, M. S. , &Williams, J. M. (1993). Erythroid

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specific antibodies enhance detection of fetal nucleater erythrocytes in maternal blood. Prenatal Diagnosis 13,

Burton, B. K., Schulz, C. J., 8r Burd, L. I. (1992). Limb anomalies associated with chorionic villus sampling. Obstetrics & Gy- necology, 79,29.

Canadian Collaborative CVS-Amniocentesis Clinical Trial Group (1989). Multicenter randomised clinical trial of chorion villus sampling and amniocentesis: first report. Lancet, 1, 2.

Canick, J . A. , & Knight, G. J . (1991). Multiple marker screening for fetal Down syndrome. Contemporary OB/GY& 3- 1 1 .

Chesheir, N . C., & Cefalo, R. C. (1992). Prenatal diagnosis and caring. Women’s Health Issues, 2(3), 123-132.

D’Alton, M. E . . & DeCherney, A. H. (1993). Prenatal diagnosis. New England.Journal ofMedicine, 328, 116-1 18.

Gray, J . W., Kallioniemi, A., Kallioniemi, O., Pallavicini, M., Waldman, F., 8r Pinkel, D. (1992). Molecular cytogenetics: diagnostic and prognostic assessment. Current Opinion in 13iotechnolo.g: 3,623-631.

293-300.

Thompson, M. W., McInnes, R. R., 8r Huntington, F. W. (1991). Geneticsin medicine (5th ed). Philadelphia: WB Saunders.

Wald, N., Cuckle, H., Lk Nanchahal, K. (1989). Amniotic fluid acetylcholinesterase measurement in the prenatal diagnosis of open neural tube defects: 2nd report of the collaborative acetylcholinesterase sudy. Prenatal Diagnosis, 3,

Ward, B. E. , Gerson, S. L. , Carelli, M. P., McGuire, N. M., Dack- owski, W. R., Weinstein, M., Sandlin, C., Warren, R., 8r Klinger, K. W. (1993). Rapid prenatal diagnosis of chromosomal aneuploidies by fluorescence in situ hybridization: Clinical experience with 4500 specimens. American Jour- nal ofHuman Genetics, 52,854-865.

813-829.

Address for correspondence: Lynette Wright, RN, MS, Rtchlyn Associates, 2391 Woodleaf Lane, Decatur, GA 30033.

Lynette Wright ts a genetic educatton consultant and president and founder of Richlyn Associates in Decatur, GA.

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