Generally, people who chose to have prenatal diagnosis (PND) are concerned about some specific chromosomal condition, the most common of which is Down syndrome in the context of older child-bearing age or of an increased-risk abnormal screening test. The major categories of unexpected chromosomal abnormality are (1) an autosomal trisomy other than trisomy 21, (2) a sex chromosome aneuploidy, (3) a structural rearrangement, (4) an extra structurally abnormal chromosome, (5) polyploidy, and (6), for each of the foregoing, mosaicism.
DECISION MAKING FOLLOWING PRENATAL DIAGNOSIS OF A CHROMOSOMAL ABNORMALITY
To some extent, the possibility of other abnormalities should be raised at counseling before PND. But when an abnormality is actually discovered, it is of course necessary to discuss in detail with the couple the implications of this particular abnormality and to help them decide on a suitable course of action. Outlines of the clinical consequences of these abnormalities follow, to serve as a basis for these decisions. In transmitting the information, the counselor is obliged to be clear and accurate about the particular abnormality and to take care that the parents' autonomy in the decision-making process is not compromised. The difficult decision for or against continuation of the pregnancy is the immediate one to be made. Murray (2003) emphasizes the usefulness of knowing about an abnormality ahead of time, in order that perinatal management may be better informed, for those for whom termination would not be acceptable.
Unsurprisingly, the severity of the condition influences decision making. Drugan et al. (1990) found that 93% of parents having a prenatal diagnosis with a poor prognosis (autosomal trisomy, unbalanced translocation, 45,X with major ultrasonographic defects) chose pregnancy termination, while only 27% of parents given a questionable prognosis (sex chromosome aneuploidy, 45,X with normal ultrasonography, de novo apparently balanced translocation or inversion) took this course. They make the interesting observation that ultrasound visualization of fetal defects “in a society dominated by the television screen” can be useful in helping parents better grasp the implications of the diagnosis.
With respect to Down syndrome (DS), views differ. A study of health professionals in Finland, for example, showed some inconsistency in the points of view of midwives and public health nurses with the options available to their patients (and the acknowledgment of this difference could be seen as a healthy sign) (Jallinoja et al., 1999). Most (79%) of these midwives and nurses agreed that all pregnant women should be offered a screening test, although only 44% personally accepted the concept of genetic abortion. An acceptance of abortion correlated with education and with a professional experience with DS patients. In the United States, Britt et al. (2000) studied 142 women who had had a PND of trisomy 21, seen in Detroit over the period 1989–1998. Those who had already had children, and where the diagnosis of trisomy 21 was made earlier in the pregnancy, were more likely to choose termination, as were childless women having a diagnosis later (after 16 weeks) in pregnancy.
The grayest area is sex chromosome aneuploidy, and views have been changing somewhat over recent decades, generally in the direction of a more conservative response to the news of a chromosomal abnormality (Christian et al., 2000; Linden et al., 2002). In Denmark in 1986, Nielsen et al. reported that approximately 80% of prenatal diagnoses of sex chromosome aneuploidy at that time were followed by the choice of abortion. In an English/ Finnish study from the same period, termination (in about 60% overall) was more likely to be chosen in the case of the XXY and 45,X karyotypes by younger parents with fewer previous children, and in all cases in which an ultra-sonographic defect was identified (Holmes-Siedle et al., 1987). From a large survey of centers in five European countries, covering the period 1986 to 1997, the rate of choice of termination with respect to XXY was 44% (Marteau et al., 2002). In a German study over a similar period, termination was chosen by a much smaller fraction, only 13%, among parents who had been given a prenatal diagnosis of 47,XXX, 47,XXY or 47,XYY (Meschede et al., 1998c). This may in part have reflected the practice of this clinic to emphasize the point that “the mean global IQ of around 90 falls well within the normal range and is compatible with a productive and socially well-adjusted life.” In contrast, just 2% of parents at the same clinic decided to continue a pregnancy with trisomy 21. Parental attributes may be important: in theexperience of the Denver group, those parents choosing prenatal diagnosis were often of higher socioeconomic status, and had made conscious decisions to continue the pregnancy following the discovery of a sex chromosome abnormality, and these children had generally done better than those identified in whole population newborn surveys (Linden and Bender, 2002).
The way in which information is given has an important impact, and counselors need to be well aware of the weight that patients, in some emotional turmoil at the news they have just received, may put upon the information. Consider the example of 47,XXY Klinefelter syndrome. In the European survey mentioned above, Marteau et al. (2002) assessed responses to the prenatal diagnosis of XXY when counseling had been given by obstetricians, pediatricians, midwives, health visitors, or genetics specialists. Women counseled solely by genetics specialists were more than twice as likely (relative risk = 2.4) to continue the pregnancy than those counseled either by other professionals or by other professionals along with a geneticist. It seems probable that these differences may reflect the style of counseling. Marteau et al. (1994) make the following distinctions: nondirective counseling (“try to be as neutral as possible, covering both positive and negative aspects”), directive counseling for termination (“encourage termination” or “try to be as neutral as possible but overall convey more negative than positive aspects of the condition”), or directive counseling against termination (“encourage parents to carry to term” or “try to be as neutral as possible but overall convey more positive than negative aspects of the condition”). A consistent approach, with access to accurate information, is to be emphasized, as is, of course, the requirement of enabling women to be well informed, in the broadest sense, to make the choice that will sit best with them (Abramsky et al., 2001; Marteau and Dormandy, 2001; Linden et al., 2002). Beyond the clinic, there are support groups, public information resources, and talking with other parents, as means to become further informed about the implications of a sex chromosome abnormality (in the short period of time during which a decision must be made), and Linden et al. (2002) note the pros and cons of taking these paths. The prime responsibility for putting couples in the best position to make an appropriate decision lies with the counselor.
MOSAICISM: CONSTITUTIONAL, CONFINED, AND PSEUDO
Mosaicism is the bane of cytogenetic prenatal diagnosis. Most times, it turns out to have been a false alarm, and the mosaicism in villus tissue or amniocytes does not reflect a true constitutional mosaicism of the embryo. This is a problem for the laboratory to resolve, not the patient. A chromosomally abnormal cell line may exist only in extraembryonic tissues (chorion, amnion), and the embryo is 46,N. This is confined placental mosaicism (CPM). CPM, is an issue arising at chorionic villus sampling (CVS) rather than at amniocentesis. Or, embryonic and extraembryonic tissues are all 46,N, and the abnormality arose during tissue culture in vitro (cultural artifact). This is pseudomosaicism (this expression is sometimes erroneously used in referring to CPM). In those rare instances when mosaicism does truly involve the embryo, we usually cannot offer, on the basis of the karyotype per se, a firm prediction of what effect this may have on the phenotype. Considerable discussion follows: but at the outset, we should emphasize that true mosaicism of the fetus is very infrequently observed, and the majority of mosaicism identified at prenatal diagnosis, more especially at CVS, does not presage an abnormal baby. It is important to keep this perspective in talking with parents (according to the particular attributes of the mosaicism, as we go on to discuss), and avoid causing any more anxiety than that which, inevitably, an “abnormal” result brings.
Applied Embryology
Interpreting mosaicism obliges an understanding of the earliest events of development of the conceptus (Bianchi et al., 1993; Robinson et al., 2002). The zygote undergoes successive mitoses to produce a ball of cells (morula) (Color Fig. 25-1, see separate color insert). The morula then cavitates to produce an inner cyst, and becomes the blastocyst (this is happening at the beginning of the second week post-conception) (Color Fig. 25-1, 2, see separate color insert). The outermost layer of the blastocyst consists of trophoblast, and this tissue becomes the outer investment of the chorionic villi. The inner cell mass protrudes into the blastocystic cavity, and it will give origin to the embryo. It comprises two different cellular layers, the epiblast and the hypoblast. In a 64-cell blastocyst, most cells are trophoblasts, the inner cell mass comprises about 16 cells, within which only about 4 (epiblast) cells will give rise to the embryo itself.
The hypoblast forms the spherical primary yolk sac (whose roof is, transiently, the ventral surface of the embryo). The primary yolk sac gives rise to the extraembryonic mesoderm, sandwiched between itself and the outer cytotrophoblast, thus producing a three-layered sphere. The mesodermal cells now invade the blastocystic cavity (Color Fig. 25-1, 3, see separate color insert), and this mesodermal mass is in turn cavitated to produce the extraembryonic celom, such that there are outer and inner layers of extraembryonic mesoderm. The outer layer, underlying the trophoblast, gives rise to the mesenchymal core of the chorionic villus, and the inner layer becomes the outer (mesodermal) surface of the amniotic membrane. The amniotic cavity enlarges at the expense of the extraembryonic celom (Color Fig. 25-1, 5, see separate color insert; Fig. 25-2), and eventually obliterates it (by the end of the first trimester), with the mesodermal layer of the amnion fusing with the mesodermal layer of the chorion.
The epiblast gives rise to the amniotic cavity, the floor of which is the “dorsal” (ectodermal) surface of the embryo, and its roof is the amnion, these being continuous at their margins. Thus, the embryonic integument and the inner surface of the amniotic membrane, which are the source of the embryonic and amniotic epithelial cells present in amniotic fluid, have the same lineage. At the beginning of the third week, the primitive streak arises from the epiblast, and this in turn gives origin to both endoderm and intraembryonic mesoderm. Thus, endoderm and intraembryonic mesoderm are closely related developmentally to ectoderm. Endoderm gives origin, among other tissues, to urinary tract and lung epithelia, desquamated cells from which contribute to the cellular population of amniotic fluid. Albeit that the extraembryonic and intraembryonic mesoderms have different origins, there may be migration of some intraembryonic mesodermal cells into the (extraembryonic) amniotic mesoderm. Cells from the latter add a minor fraction to the population of amniocytes, but have a proliferative advantage, and may come to comprise most of the cells present following in vitro culture.
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Figure 25-2. Ultrasound picture of embryo at 10–11 weeks gestational age, quite close to actual size (note cm. markers at right). Note amnion (A), amniotic cavity (AC), extraembryonic celom (EC), umbilical cord (U), “physiological omphalocele” (O), yolk sac (Y), and placenta (P). (Courtesy H. P. Robinson.) The relative positions of embryo and other structures are similar to the depiction in the drawing in Color Figure 25-1 part 5. |
Amniocentesis is, therefore, a procedure that samples cells having origin from the epiblast of the inner cell mass, and these cells rather closely reflect the true constitution of the embryo. Chorionic villus sampling, by contrast, samples more distantly related cells: trophoblast cells (direct and short-term culture), which were the first lineage to differentiate from totipotent cells of the morula, and villus core cells (long-term culture), which reflect the more recently separated lineage of the extraembryonic mesoderm. The differing origins of tissues sampled by different means are set out in Figure 25-3.
Mechanisms of Mosaicism
Mosaicism may involve aneuploidy for an intact chromosome or for an abnormal chromosome, along with a normal cell line. Mosaic trisomy can arise by one of the mechanisms set out in Figure 2-8. The distribution of the normal and the abnormal cell lines in the fetus and the placenta depends upon the time and the place of the abnormal mitotic event. If a trisomic conceptus is rescued at a very early stage, in a cell that is going to give rise to the inner cell mass and to some of the extrafetal tissues, then the embryo may be 46,N, and the placenta will show mosaic trisomy. If rescue occurred at a later stage, the placenta might be entirely trisomic, with a mosaic trisomy of the fetus. These and other possible combinations are depicted in Figure 25-4.
The eventual phenotype will be influenced by the tissue distribution of the cell lineages that contain the trisomic chromosome and the normal/trisomic proportions in various tissues. If, say, a nondisjunction occurs in 1 of the 50 or so trophoblast cells of a 46,N 64-cell blastocyst, the lineage of this one cell may contribute trisomic tissue to the placenta, but the fetus (deriving from the inner cell mass) will be 46,N. Trophoblast cells are very rapidly dividing, for the crucial task of providing tissue to make an attachment to the uterine wall (which happens at the beginning of the second week), and this may endow a greater vulnerability to mitotic error. Certain other factors may, theoretically, predispose to the generation of early post-conception mosaic aneuploidy (Wolstenholme, 1996).
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Figure 25-3. Diagram of cell lineages arising from differentiation in the very early conceptus. The fertilized egg (1) produces a trophoblast precursor (1b) and a totipotent stem cell (2) which in turn forms another trophoblast precursor (2b) and a stem cell (3) that produces the inner cell mass. The inner cell mass divides into stem cells for hypoblast (3b) and epiblast (4). The epiblast cell(s) (5) produce embryonic ectoderm and primitive streak, and the latter is the source of embryonic mesoderm and endoderm. The cell lineages sampled at various prenatal diagnostic procedures are indicated at right. E, epiblast; H, hypoblast; P, primitive streak; Y, yolk sac. (From Bianchi, D. W., Wilkins-Haug, L. E., Enders, A. C. and Hay, E. D. (1993), Origin of extra-embryonic mesoderm in experimental animals: relevance to chorionic mosaicism in humans. Am. J. Med. Genet. 46, 542–550, © 1993 Am. J. Med. Genet., courtesy D. W Bianchi, and with the permission of Wiley-Liss, Inc., a subsidiary of John Wiley & Sons, Inc.) This construction is to be compared with that of Kennerknecht et al. (1993b), in which three postzygotic mitoses occur, producing eight totipotent cells, before the cells begin to take on their tissue identities. Robinson et al. (2002) propose a further variation, with some cells of the embryonic mesoderm migrating into the (otherwise extraembryonic) mesodermal layer of the amnion. |
The potential for widely differing tissue distributions of the different cell lines may confound interpretation at PND. Consider the case of Jewell et al. (1992). A dup(12) chromosome was present in 87% of amnion cells, 60% of fetal blood, but only 2% of chorionic villi and in 0% of chorionic membrane. Kingston et al. (1993) provided a similar remarkable (and disconcerting) example. Amniotic fluid cells had 3% with an extra structurally abnormal chromosome (ESAC), a sample of fetal blood showed all cells to be 46,N, and several tissues taken post-termination had various fractions of mosaicism, including brain with 88% of cells aneuploid. Stankiewicz et al. (2001c) report a nonmosaic 46,X,der(Y)t(Y;7)(p11.32; p15.3) causing a 7p trisomy syndrome in an infant, following a CVS diagnosis of very low-grade mosaicism 46,X,der(Y)t(Y;7) [1]/46,XY[49], and yet with nonmosaic 46, X,der(Y)t(Y;7) at amniocentesis. These observations point to an early postzygotic origin of the translocation in an initially 46,XY conceptus, apparently affecting the entire inner cell mass but only a very small minority of trophoblasts. These three cases, admittedly exceptional, are instructive in emphasizing that the proportions of abnormal cells in one tissue can not necessarily be taken as indicative of proportions elsewhere.
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Figure 25-4. Types of mosaicism of the fetal–placental unit. Fetus is depicted enclosed in its sac at right, with the chorionic villi comprising the placenta to left. Gray areas indicate an aneuploid cell line; white areas indicate karyotypic normality. In reality, the distributions of the two cell lines are unlikely to be as clearcut as is shown here. In the examples showing placental mosaicism, the path taken by the sampling needle will determine whether the abnormality is detected or missed at chorionic villus sampling. The cartoon of the fetus, sac, and placenta is close to the form and about two-thirds the size that actually exists at 10 weeks 0 days (gestational age as measured clinically, dated from the last menstrual period), when crown–rump length is around 30 mm. |
Laboratory Assessment of Mosaicism
The resolution of mosaicism in the cytogenetics laboratory and in its clinical interpretation can differ for CVS and amniocentesis, and we will consider them separately. In terms of the laboratory result, we can apply to both CVS and amniocentesis the concept of different levels of in vitro mosaicism, originally developed for amniocentesis by Worton and Stern (1984) and refined by Hsu et al. (1992) and Hsu and Benn (1999), as follows.
Level I. A single abnormal cell is seen. With near certainty this is cultural artifact, and is thus pseudomosaicism, and this may be resolved as set out in Tables 25-1 and 25-2. The laboratory would not usually report the observation.2
Level II. Two or more cells with the same chromosomal abnormality in a dispersed culture from a single flask are seen, or in a single abnormal colony from an in situ culture (i.e., possibly or probably just a single clone). Some would also include the observation of two or more colonies from the same in situ culture. The abnormality is not observed in colonies from other independent cultures. This form of mosaicism is almost always pseudomosaicism. It would not usually be reported to the physician, but it may be if additional studies are inadequate, if fetal anomalies were identified, or in the case of certain chromosome abnormalities that are well recognized as existing in the mosaic state. A course of action to resolve the issue cytogenetically, in the case of amniocentesis, is given in Table 25-1.
Level III. Two or more cells with the same chromosome abnormality are distributed over two or more independent cultures. Level III is likely to reflect a true mosaicism, and the cytogeneticist will report this finding immediately. (Some allow level III to include more than one colony in only a single flask, although this could be an “overinterpreted level II” if two colonies in the one flask had arisen from a single cell whose progeny migrated and established separated clones.)
The distinction may not be quite as clear as this in practice, but this is a useful working definition.
The mathematics of sampling comes into the picture: how many cells need to be looked at, to establish a level of confidence that mosaicism of a particular extent can safely be disregarded? Tables have been derived to assist in answering this question (Hook 1977; Sikkema-Raddatz et al., 1997a). Inevitably, mosaicism will, on rare occasions, be missed. Given the reality that only a limited number of cells can be karyotyped, the statistics will sometimes conspire against the cytogeneticist, and only normal cells will be examined. This has to be accepted: the test is not perfect. For example, de Pater et al. (2003a)describe their experience in reporting a normal result from amniocentesis, but in due course the child proving to be a r(12) mosaic, 50% on blood. Critically reviewing their procedures, and indeed being able to see the ring chromosome when archived material from the amniocentesis was restudied, they nevertheless drew the conclusion that their original analysis had been appropriately performed. A similar example, with respect to a CVS case, is noted below, in the section on 47,i(5p). In CVS, the exposure to error may relate to the part of the placenta the sampling needle happens to traverse.
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Table 25.1. Guidelines for Work-Up in Elucidation of Possible Amniocyte Pseudomosaicism/Mosaicism |
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Table 25.2. Types and Frequencies of Placental–Fetal Discordance at Chorionic Villus Sampling |
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Prediction of Phenotype in an Individual Case
Elegant theorizing notwithstanding, the pragmatic observations from published cases in the literature provide the mainstay of the advice that the counselor may offer the parents in an individual case. (Large series are better than single case reports, which are better than anecdote.) Are mosaicisms for some particular chromosomes, or types of aberration, of more concern than others? What is a low enough level of mosaicism, if any one exists, to have a degree of confidence that the child will be normal? We set out below summaries of the recorded examples from the literature, none of which necessarily provides a firm answer, but which may serve as the basis for discussion and counseling. The numbers in some are very small. Another difficulty with these data is that, for the most part, the window of observation of the child's phenotype was confined to the neonatal period. Of course, many children who are eventually diagnosed with significant handicap may have been well grown and morphologically normal at birth and with normal functional neurology (inasmuch as this may be assessed in a baby). It is possible, however, to overdiagnose problems in babyhood, as a child who subsequently develops normally may prove (Warburton, 1991; Joyce et al., 2001). An important concern in mosaicism is that a cell line inaccessible to analysis—specifically, in the brain—might contain the abnormal chromosome, despite a normal karyotype in the postnatal tissues that are normally examined, namely, blood and possibly skin. If so, cognitive functioning could be compromised. Those few reports that include follow-up data for some years into childhood (8- and 9-year-olds from prenatal diagnoses of trisomy 20 mosaicism studied in Baty et al. [2001] being shining examples) are therefore most valuable. Nevertheless, no certainty can be offered, recognizing that every case of mosaicism will be unique, in terms of the extent and qualitative tissue distribution of the abnormal lineage.
CHORION VILLUS CULTURE AND MOSAICISM
Chronic villus sampling mosaicism is detected in 1%–2% of procedures at the 10- to 11-week mark. Mosaicism from an early mitotic error can give rise to confined mosaicism (confined to placenta, or to fetus) or to generalized mosaicism (present in both fetus and placenta); the broad range of possibilities is shown in Figure 25-4. The extreme form is complete discordance, with a nonmosaic 46,N karyotype in fetus and nonmosaic aneuploidy in CVS, or vice versa. Depending on the timing and site of the event or events producing the mosaic state, the karyotypes observed at short- and long-term CVS will vary. Six scenarios for placental–fetal mosaicism are set out as types I–VI in Table 25-2 (Hahnemann and Vejerslev, 1997). For each of these, in theory at least, two formats may apply: first, a mitotic error in an initially normal conceptus which gives rise to an abnormal cell line, or second, an initially abnormal conceptus, typically due to a meiotic error, with a subsequent postzygotic event generating a normal cell line.
Mosaicism detected in cytotrophoblast but not in stroma will usually be confined placental mosaicism (type I). In the case of mosaic trisomy, the risk for uniparental disomy (UPD) due to postzygotic “correction” may, however, be high (Wolstenholme, 1996). (Some laboratories have ceased routine direct and short-term cultures, and thus for them a question of type I CPM no longer arises.) Mosaicism identified in stroma may be confined (types II and III) or generalized (types V and VI); making the distinction between these, and from in vitro artifact, is not necessarily straightforward. It may be appropriate to follow the process outlined for amniotic fluid culture in Table 25-1 for the flask method to help resolve level II mosaicism at CVS, but it is almost always the case that the fetus/baby turns out to be chromosomally normal (Ledbetter et al., 1992; Fryburg et al., 1993). In contrast, level III mosaicism (discussed in detail below) very probably reflects a true mosaicism, at least of the placenta, and potentially of the fetus.
False-Negative Results
The important distinction to make here is between direct (uncultured) and cultured samples. In a review of reports up to 1997, Sikkema-Raddatz et al. (1997b) collected 20 examples of fetuses misdiagnosed as normal from CVS culture. Seven cases involved a sex chromosome mosaicism of the fetus (45,X/46,XX; 45,X/46,XY; 47,XXX/46,XX; 47,XXY/46,XY; 47,XYY/46,XY), and in 13 there was a fetal trisomy 18 or 21, mosaic or non-mosaic. In the majority, a direct analysis had given a 46,N result, with a CVS culture going on to demonstrate an abnormal karyotype, which was subsequently confirmed at amniocentesis, fetal tissue analysis, or karyotyping of the infant. These reflected a type V placental–fetal mosaicism. In five, a CVS culture was not done, and the abnormality was not identified until amniocentesis or fetal/neonatal studies had been undertaken. Only one case showed a normal karyotype on both direct and cultured CVS, with the child having trisomy 18 (this type not accounted for in Table 25-2) (Pindar et al., 1992).3 This statistic is to be emphasized: in the entire world literature, just one single case reported of a false negative result from a cultured CVS analysis. Thus, practically all of the time, a normal long-term CVS result means that the baby will be chromosomally normal. Other possible theoretical routes whereby a true mosaicism might be missed at CVS include a pregnancy in which there had been a resorbed 46,N twin, with the sampling instrument having passed through its 46,N placental trace; statistical misfortune when sampling of metaphases at microscopy fails to find the abnormal cell line; and contamination with 46,XX maternal cells.
Origin of Mosaic Trisomy
Robinson et al. (1997) studied 101 cases in which CPM had been identified at CVS, seeking to establish correlates of the origin of the trisomy. Some CPM trisomies are usually of mitotic (somatic) origin, the zygote having been 46,N. Others typically arise meiotically, and the zygote was trisomic ab initio. That is to say, meiotic or mitotic origins of the trisomy are substantially chromosome-specific. For example, almost all cases of CPM for trisomy 16 have arisen at maternal meiosis I, while in contrast, trisomy 8 CPM is characteristically the consequence of a mitotic event. In the trisomy 16 cases, UPD 16 could be identified in 18%; in all cases, this was a maternal UPD, and the placental trisomy had been of meiotic origin. In other words, every case of fetal UPD 16 had arisen as the consequence of “correction” of an initially trisomic conceptus, which in turn was due to a maternal nondisjunctional event. As Robinson et al.emphasize, their study population was not a random sample of mosaic cases, with some ascertained through ultrasonographic abnormality. But it is certainly notable that all but one of the intrauterine growth retardation (IUGR) cases in their survey reflected a meiotic origin of the CPM. As for CPM of somatic origin, the frequency of somatically arising errors in their series may, for various reasons, have been underestimated. Somatic mosaicism may certainly be associated with fetal normality, but it is often the case that the trisomic fractions detected at CVS are small. Trisomy 2 at CVS is an example of a mosaicism that conveys quite different implications according the meiotic or mitotic mechanism of its generation (see below).
Level III Mosaicism
Level III mosaicism raises an immediate concern. Management at this point (which will usually be around 12 weeks) is aimed at demonstrating, as much as possible, fetal normality, or, if it transpires, confirming a true fetal mosaicism. Amniocentesis with rapid FISH analysis of a large number of cells, along with detailed ultrasonographic assessment of fetal morphology, is usually called for. In fact, the majority will return normal results.
A large amount of data on level III mosaicism for autosomal trisomy was gathered by the European Collaborative Research Group on Mosaicism in CVS (EUCROMIC) (Hahnemann and Vejerslev, 1997), comprising information on just over 92,000 CVS procedures from 79 laboratories during 1986–1994. Mosaicism (or nonmosaic fetoplacental discrepancy) was seen in 650 (1.5%) cases. Of these, 192 were followed up in detail, with karyotyping of fetal fibroblasts, fetal blood, amniocytes, or neonatal tissues. Most, 84% of the 192, represented CPM. The abnormal cell line was present in either trophoblast (50%), villus mesenchyme (30%), or both (20%). A similar proportion was forthcoming from another large review, that of Phillips et al. (1996), comprising 469 cases of placental mosaicism identified at CVS in 13 separate studies, in only 50 (11%) of which was fetal mosaicism actually demonstrated. A greater risk applied when the abnormality had been detected on villus culture and when the chromosome concerned was a marker or one of those involved in the common trisomies. These authors emphasized the value and validity of follow-up amniocentesis.
Certain CVS trisomies are more or less likely to reflect the same trisomy in the fetus, and the pattern and distribution of the cell lines are also indicative, as set out in Table 25-3. Trisomy 21 mosaicism on CVS is the most likely to represent a true fetal trisomy 21, whether in the non-mosaic or mosaic state. A risk applies also with trisomies 8, 9, 12, 13, 15, 18, and 20. On the other hand, CPM or fetoplacental discrepancy for trisomies 2, 3, 5, 7, 10, 11, 14, 16, 17, and 22 was never, in the EUCROMIC series, confirmed at fetal or postnatal studies. In some trisomies, a true fetal mosaicism may exist, but at such a low level that there might be no discernible effect upon the phenotype. Klein et al. (1994) reported such a case, a child born of a pregnancy in which trisomy 8 was observed in 81% of CVS cultured cells, 0% of amniotic fluid cells, and in 60% of a placental biopsy at delivery: the child had 4% and 1% mosaicism in blood at 2 and 7 months of age and 0% on a skin fibroblast study, and was normal in appearance, growth, and developmental progress at age 30 months. Of course, fetal morphologic defect shown on ultrasonography would indicate the very substantial probability of a major degree of true fetal mosaicism, and in that case the choice of termination is appropriately offered.
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Table 25.3. Likelihood of a Fetal Trisomy (Whether Mosaic or Nonmosaic) According to Distribution of Trisomic Cell Line in Chorionic Villus Sampling |
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The vagaries of sampling may influence the interpretation, as the following examples show. We followed to term a woman in whom first-trimester CVS had shown trisomy 7 mosaicism with 47,XY,+7 in three out of eight clones; and yet three out of four placental samples, and peripheral blood from the (normal) baby, karyotyped 46,XY. Just one placental sample, which was not histologically distinguishable from the others, was 47,+7 (Watt et al., 1991). It may well have been that the CVS sampling catheter had traversed this unrepresentative region of the placenta, and most of the sample that was eventually analyzed came from here. Similarly, in a case of i(5p) diagnosis at CVS (see below, Autosomal Isochromosome), following the birth of the (normal) baby, we identified a region of placental mosaicism (Clement Wilson et al., 2002). De Pater et al. (1997) did a CVS in a pregnancy of 37 weeks gestation in which severe growth retardation and a heart defect had been identified, and this showed nonmosaic trisomy 22. However, from a simultaneous amniocentesis, only 2 out of 10 clones were 47,XX,+22, the other 8 being normal; and a cord blood from the (abnormal) baby gave a nonmosaic 46,XX karyotype. Skin fibroblasts demonstrated mosaicism, 47,XX,+22[7]/46,XX[25]. Of 14 placental biopsies studied by interphase FISH, only one showed trisomy 22 cells, and at a low (about 20%) percentage. Again, it may be that a small nidus of trisomic tissue happened to be in the path of the CVS sampling needle, and the sample was aspirated while the needle was at this very spot. (This is an example of fetal–placen-tal mosaicism, as illustrated in Fig. 25-4.)
A specific concern when CPM for a trisomy is diagnosed relates to UPD. This is an issue in those trisomies involving an imprintable chromosome (including 7, 11, 14, 15, and possibly 16). The embryo may “correct” by postzygotic loss of the additional chromosome, while the placenta remains partly or wholly trisomic. In the particular case of 47,+15/46 or 47,+15 diagnosed at CVS, and if a 46,N karyotype is shown at the subsequent amniocentesis, a Prader-Willi/Angelman methylation test can be applied to the cultured amniotic fluid cells. Other UPDs seem mostly to be without phenotypic effect per se (Jones et al., 1995; Soler et al., 2000).1 Irrespective of imprinting, there remains also the question of a small residual trisomic cell line in the fetus, potentially contributing to an abnormal phenotype.
Effect on Placental Function
If a cytogenetically abnormal cell line is confined to the placenta, does this have any implication for placental function? Apparently, at least for several trisomies, a placenta that is in part trisomic retains nevertheless a sufficient or nearly sufficient level of function, and as a general rule the (46,N) fetus is satisfactorily supported. Lestou et al. (2000) analyzed a series of 100 placentas from pregnancies producing a “viable and nonmalformed” infant, using the methodology of comparative genomic hybridization (CGH) with confirmatory FISH, and found one with CPM in only trophoblast (trisomy 13), two with CPM in the stroma (trisomies 2, 12), and two with mosaicism in both compartments (trisomies 4, 18). The more commonly observed CPMs at prenatal diagnosis involve chromosomes 2, 3, 7, and 8 (which mostly arise mitotically), and chromosomes 16 and 22 (mostly of meiotic origin, and typically affecting both trophoblast and villus core placental constituent parts). It is mostly CPM of meiotic origin that is associated with a risk for pregnancy complication (Kalousek, 2000).
AMNIOTIC FLUID CELL CULTURE AND MOSAICISM
A mitotic error in epiblast may produce mosaicism of both embryonic and amniotic tissue. A mitotic error in extraembryonic epithelium causes mosaicism confined to the amniotic membrane. An in vitro cell division defect causes pseudomosaicism. Separating confined placental mosaicism and pseudomosaicism from true mosaicism is critical, but by no means straightforward. The distinction is, in the first instance, based on the number of abnormal cells seen, and whether one or more than one presumptive abnormal clone exists, according to the three levels I–III set out above. Level I mosaicism is seen in 2.5%–7% of amniocenteses, level II in 0.7%–1.1%, and level III in about 2 per thousand amniotic fluids (Wilson et al., 1989).
Once the laboratory studies are completed, the cytogeneticist will provide an opinion about the level of mosaicism, taking into account technical aspects of the cultures. There is generally no point, and indeed it could be counterproductive, to report level I mosaicism. Some level II mosaicism and all level III mosaicism, however, must be conveyed to the patient, and be carefully and clearly interpreted.
Level II Mosaicism
Level II mosaicism reflects a true fetal chromosomal abnormality in only 1% or less of cases (Worton and Stern, 1984; Ledbetter et al., 1992; Fryburg et al., 1993; Liou et al., 1993). The nature of the “mosaic chromosome” is important. If it is one that has been recorded, in life, in the nonmosaic trisomic state, or in the mosaic state, the level of concern is higher. This includes, for example, mosaic trisomies 8, 9, 13, 18, and 21 and mosaic isochromosomes 5p, 9p, 12p, and 18p. Although true mosaicism for most of the trisomies has been observed in the malformed fetus in a pregnancy advancing well into the third trimester or in an abnormal liveborn child, these cases are so rare that a level II amniotic fluid mosaicism is still more likely due to artifact than a true significant fetal mosaicism. High-resolution ultrasonography can provide helpful information in this context.
If further cytogenetic investigation is judged desirable, and it often is, repeat amniocentesis for interphase FISH analysis is the procedure of choice, with probe choice according to the chromosome in question. A large number of cells can be analyzed, and quickly. Fetal blood sampling, formerly the mainstay, is much less used nowadays. It is to be noted that not all mosaicism is necessarily present in blood, and, for example, fetal blood sampling only infrequently (if ever) detects a mosaic cell line in trisomy 5, 12, 20, or 22, or i(12p) (Berghella et al., 1998; Chiesa et al., 1998).
Strictly speaking, no amount of investigation could ever completely exclude the possibility of a true mosaicism of the fetus, even if the distribution of the abnormal cell line may be rather limited and of unimportant phenotypic consequence. We have seen, for example, a case of level III 47,XX,+13/46,XX mosaicism at CVS, followed by the demonstration of very low-level mosaicism at amniocentesis (1/28 colonies trisomic) and fetal blood sampling (1/400 cells trisomic). At birth, a cord blood sample from the baby showed 47,XX,+13 in 1 out of 150 cells; 2/32 cells were trisomic in amnion and 1/30 and 3/30 in two placental villus biopsies (Delatycki et al., 1998). It only needed the colony from one amniocyte not to have been analyzable or one lymphocyte to have been passed over at each blood sampling for the true state to go unrecognized. The child has since developed normally (M. B. Delatycki, pers. comm., 2001). Rare similar examples exist to disquiet the counselor (Terzoli et al., 1990; Vockley et al., 1991), but a sense of perspective is to be kept: for each autosome, only the tiniest number of level II mosaicisms (zero for most chromosomes) have turned out to reflect, in fact, a recognized true mosaicism of the fetus.
Level III Mosaicism
Hsu and Benn reevaluated the issues in 1999, and they have set forth useful guidelines. These are presented in detail in Table 25-1. While every autosome has now had a mention as a mosaic trisomy at prenatal or postnatal diagnosis, some are very rare, and others are of questionable significance. Some reported associations may not necessarily have been causal. Hsu and Benn propose the stringent requirement that, before embarking on an extensive work-up, for the particular chromosome there should be in the literature “two, or more, well-documented independent reports of confirmed amniocyte mosaicism with abnormal pregnancy outcomes.” The most extensive data treating the question are published in two reports from a collaboration of a number of American and Canadian laboratories: Hsu et al. (1997) with respect to the rare trisomies, and Wallerstein et al. (2000) on trisomies 13, 18, 20 and 21. We make much use of this material in the commentaries below, and every PND laboratory will want to have a copy of these papers readily at hand. Ultrasonography provides useful adjunctive evidence, but apparent normality cannot be taken as a guarantee. Studies for UPD may need to be considered in the case of mosaicism for chromosomes known to be subject to imprinting. Further modifications to these guidelines can be anticipated, as new data come to hand.
One should always attempt to confirm a diagnosis of mosaicism, either on multiple fetal samples following pregnancy termination, or on blood and placenta in an infant. A post-termination study that did not confirm the abnormality could cause parents great distress, and it should be decided with them beforehand whether they would be given the result. An unconfirmed abnormality could be misleading in a twin pregnancy in which the diagnostic sample came from a vanishing abnormal twin, but the post-termination tissue came from the normal co-twin (Griffiths et al., 1996). Fejgin et al. (1997) refer to the “hopeful possibility” of mosaicism as a comfort to parents, with the post-termination tissue having sampled only the normal cell line. It is true that even multiple tissue sampling cannot be taken as having ruled out mosaicism, and a diagnosis of “apparent phenotypic normality” in a fetus still leaves open that a functional brain defect could have come to pass.
SPECIFIC ABNORMALITIES
In this section we attempt to outline the risks for phenotypic abnormality of specific chromosomal abnormalities detected at PND. Since the available data often derive from terminated pregnancies in which only major anomalies are recognized, many of these risk figures may be underestimates. For example, a trisomy 21 fetus may appear normal externally, but we naturally assume mental defect would have resulted; the same may apply to several other chromosomal imbalances. New knowledge will continue to accumulate, and what appears here is written on paper, not in stone.
The small number of aneuploidies that may exist in the true nonmosaic state are noted first. In the mosaic list, almost every chromosome is represented, although in the CVS section we do include also a few instances of nonmosaicism. In these, if the pregnancy would seem to be viable, it is rather probable that there would be a mosaic state in the fetus, on the understanding that these trisomies are typically lethal and would have led to miscarriage if truly nonmosaic.
AUTOSOMAL TRISOMY, NONMOSAIC
Trisomies 13 and 18 (and rarely 8, 9 and 22) are practically the only nonmosaic autosomal trisomies besides +21 that are detected at amniocentesis. Others occur but virtually all miscarry before the usual time of amniocentesis. Chorion villus sampling by contrast, is done at a gestational stage when a number of trisomies destined to abort have not yet done so. Some CVS trisomies may be nonmosaic on that tissue, but in fact the fetus and amniotic membranes are mosaic (or very rarely, even normal) (Fig. 25-4).
Trisomies 13 and 18
There is a high likelihood (43% for +13 and 68% for +18%; Hook, 1983) of spontaneous abortion after amniocentesis, and presumably it is somewhat higher if detection is by CVS. But the outlook for a liveborn child is so bleak, with inevitable profound mental deficiency, barely a vestige of social response in those few who survive beyond early infancy, and typically a requirement for full nursing care, that termination is sought by the majority of couples (Smith et al., 1989; Van Dyke and Allen, 1990; Tunca et al., 2001). Those who decide to maintain the pregnancy should know of the high perinatal and early infant mortality, the high likelihood of congenital malformation and the rarity (but not impossibility) of survival beyond infancy (Brewer et al., 2002; Kelly et al., 2002). Many would regard life-sustaining emergency surgery to the newborn as inappropriate (Bos et al., 1992). Carey (2001) puts forward a softer view, and emphasizes the need to bring the parents fully into the making of any decisions.
Trisomy 21
We expect most readers will have an expert appreciation of the predicted Down syndrome (DS) phenotype, but we do recommend Hunter's (2001) review as a full and balanced account. Marteau et al. (1994) appraised the views of obstetricians, geneticists, and genetic nurses to the prenatal diagnosis of DS, and recorded some striking differences. The respective proportions who would counsel nondirectively (see definitions above) were 32%, 57%, and 94%, and the respective proportions counseling directively in favor of termination were 62%, 40%, and 7%. About 6% of obstetricians would counsel directively in favor of continuing the pregnancy, but practically no geneticists or genetic nurses would.
Having received a positive 47,+21 result, what personal factors influence the parental decision? A 7½-year study, over 1989–1997, reports the views of 145 women in Michigan (Kramer et al., 1998). Most (87%) elected to terminate the pregnancy. The decision did not differ according to parity, race, religion, nor, perhaps surprisingly, with the presence or absence of ultrasonographic abnormality. Older mothers, those who had already had children, and those whose prenatal procedure was done at an earlier gestation were more likely to choose termination. A point to be aware of is that, with modern management, the survival of DS individuals approaches that of the general population, but comorbidities become prevalent with age, raising questions of practicalities of care as the parents themselves age (Glasson et al., 2002). On the other hand, if fetal ultrasonography shows a heart malformation and/or growth retardation, fetal death in utero or postnatal death is probable (Wessels et al., 2003).
Other Autosomal Trisomy
Never (almost) do other nonmosaic true fetal trisomies survive through to a stage of extrauterine viability. Schinzel (2001) catalogs no more than about two dozen each of trisomy 9 and trisomy 22 and barely one or two of possible trisomies 7, 8, and 14, with survival through to the third trimester. Miscarriage is nigh on inevitable, usually within the 8- to 14-week gestation range. If natural abortion has not already occurred by the time the chromosomal result is received, and if there is supportive evidence otherwise (ultrasonography, further prenatal karyotyping investigations) for there being a true fetal involvement, termination is appropriately offered. Of all the other nonmosaic trisomies, it is only with trisomy 22 that there might there be, extremely rarely, the possibility of limited postnatal survival (Tinkle et al., 2003).
AUTOSOMAL TRISOMY, MOSAIC4
Detection at Chorionic Villus Culture (in Some Cases Followed by Karyotyping at Amniocentesis)
The general rule that Robinson et al. (1997) advance is this: CVS mosaicism due to a pre-conceptual (meiotic) error conveys a significant risk for fetal trisomy/UPD, whereas a postconceptual (somatic) error is usually innocuous. Mosaic trisomies 15, 16 and 22 are mostly in the former category, while trisomies 3 and 7 are typically of mitotic origin, and mosaic trisomy 2 can be either. A somatic error is somewhat more likely when the level of trisomic cells is low, and when just one cell lineage is involved (CPM type I or II, Table 25-2). No case of UPD was associated with a mitotic origin in Robinson et al.'s study. These authors suggest that, in due course, it may become practicable and useful to develop a molecular screen to determine meiotic vs. mitotic origin to enable more precise advice about pregnancy outcome in the setting of a mosaic karyotype at PND.
The substantial majority of mosaic trisomies for a single autosome are followed by a normal result at amniocentesis and at karyotyping of the child (or of the aborted fetus). In the EUCROMIC study, there were 192 gestations with mosaic or nonmosaic fetoplacental discrepancy for an autosomal trisomy, and in 84% CPM was confirmed. For mosaic trisomy 8, 9, 12, 15 and 20, only a single case of each was subsequently identified with aneuploidy in the fetus/child, compared with two for chromosome 13, four for 18, and as many as nine for trisomy 21 (Hahneman and Vejerslev, 1997). With respect to mosaicism for multiple (>1) autosomal trisomies, the presence or absence of a normal cell line is the key point: a fetal involvement is practically never seen if there is a normal cell line, and practically always if there is no normal cell line (M. D. Pertile, pers. comm., 2002).
Most CVS mosaic trisomies “self-correct” in the sense that a follow-up amniocentesis gives a normal 46,N result. Such a result implies a chromosomally normal fetus, with a region of the placenta having been trisomic. The possibility remains for a residual effect due to (1) undetected (and presumably low level) mosaic trisomy of the fetus; (2) UPD of the fetus; and (3) placental dysfunction as a consequence ofa regional placental trisomy. The risks for these scenarios differ for different chromosomes, and we provide specific commentaries following. A rich source of information is the United Kingdom Association of Clinical Cytogeneticists database, in which are assembled the results and findings from practically every U.K. CVS laboratory over the period 1987–2000. This material is being prepared at present writing, and publication is planned (S. Connors, J. Emslie, J. Wolstenholme, pers. comm., 2003).
Mosaic Trisomy 2 at Chorionic Villus Sampling
Two broad groups of trisomy 2 mosaicism are recognized (Robinson et al., 1997; Albrecht et al., 2001; Wolstenholme et al., 2001b). In the first, a majority (85%–90% of all cases) is characterized by a small fraction of trisomic cells, and usually seen only in cultured mesenchymal cells. The pregnancy outcome is typically normal; in the series of Sago et al. (1997), 11/11 newborn infants were normal. It may be that these cases reflect a postzygotic generation of the trisomic lineage in a restricted region of chorionic tissue in an otherwise normal conceptus, and this small trisomic region has no discernible effect upon placental function. The second, minority group is presumed due to trisomy “correction” in a 47,+2 conceptus, from either a maternal or paternal error. The level of trisomic cells in the CVS is typically high, up to 100%, with the involvement of both trophoblast and the mesenchymal core. The placenta being substantially trisomic apparently compromises its function, and IUGR is a frequent observation; a residual fetal trisomy may also be a contributor. The case against UPD is yet to be settled.
Mosaic Trisomy 3 at Chorionic Villus Sampling
In the EUCROMIC study, of 10 cases with trisomy 3 at either short or long-term culture, none proved to have fetal involvement apart from one child with a normal karyotype at amniocentesis and a very low (1/100) trisomy 3 count on blood as a newborn (Hahnemann and Vejerslev, 1997).
Mosaic Trisomy 4 at Chorionic Villus Sampling
This is very rare; there were none in the EUCROMIC study. The only two recorded cases are those in Kuchinka et al. (2001). In one case, subsequent amniocentesis gave a 46,XX karyotype, but fetal demise occurred at 30 weeks, associated with considerable growth retardation (although no externally observable malformations). Upd(4)mat was demonstrated. It remains open whether the unfortunate outcome was the consequence of the UPD, or due to placental trisomy. The second case did not proceed to amniocentesis; biparental disomy 4 was demonstrated in the child. Follow-up at 1 year raised some reservation: although development was judged to be normal, growth indices were low, including a head circumference at about the 3rd centile (in other words, borderline microcephaly). To complicate the story, mother and child carried a balanced t(10;15).
Mosaic Trisomy 5 at Chorionic Villus Sampling
Only three cases are recorded in the EUCROMIC study; in none was a fetal trisomy subsequently shown (Hahnemann and Vejerslev, 1997).
Mosaic Trisomy 6 at Chorionic Villus Sampling
Very few examples are known. A detailed case report is given in Miller et al. (2001). A young mother had a 12-week CVS because of ultra-sonographic anomalies (crown–rump length at 11-week size, nuchal translucency), with 60% of cells in short-term culture and 22% of long-term cells showing 47,XX,+6. Amniocentesis was declined. An abnormal heart rate at 25 weeks led to emergency delivery, and a growth-retarded infant with numerous anomalies was born. Her blood karyotype was normal, but trisomy 6 cells were found in placenta and umbilical cord samples. Growth indices remained below the third centile. On follow-up at age 2¾ years, neurodevelopmental progress was “near normal.” Skin taken at the time of surgery showed 3% (hand) and 20% (inguinal area) mosaicism. The only two other cases on record involved mosaicism on direct preparations, followed by termination in one, and an apparently normal child subsequently born in the other.
Mosaic Trisomy 7 at Chorionic Villus Sampling
This is typically a mitotically arising mosaicism. Kalousek et al. (1997) looked at 14 cases of trisomy 7 CVS mosaicism and fetoplacental discordance, the fraction of trisomy ranging from 7% to 88% in 11, and with 3 showing 100%. Twelve infants were judged normal, and in the eight of these tested, all proved to have biparental inheritance. Two infants were of low birth weight, and the one of these tested was the only of the series with UPD and a meiotic origin; the cultured CVS in this case was 100% trisomic. In the EUCROMIC study, of 32 cases with trisomy at either or both short- and long-term culture (including three with nonmosaic trisomy), none proved to have fetal involvement (Hahnemann and Vejerslev, 1997). The conclusion is that the great majority of trisomy 7 mosaicism arises mitotically, is confined to the placenta, and does not obviously compromise intrauterine growth.
Mosaic Trisomy 8 at Chorionic Villus Sampling
A well-recognized postnatal phenotype (Warkany syndrome) accompanies trisomy 8 mosaicism, which may also include an increased risk for cancer (Seghezzi et al., 1996). Fetal defects are recorded on pathology examination (Jay et al., 1999). Typically, the mosaicism is the consequence of a postzygotic nondisjunction in an initially 46,N conceptus (Danesino et al., 1998). Van Haelst et al. (2001) reviewed their experience over the period 1986–2000, based on 33,870 prenatal tests, among which were six cases of trisomy 8 mosaicism diagnosed at CVS. These six CVS cases, as it transpired, each reflected a confined placental mosaicism, and from the five pregnancies continuing a normal baby was born. A seventh case had been reported as 46,XY normal on short-term CVS culture, but the abnormal baby had mosaic trisomy 8, thus a false-negative diagnosis. This circumstance calls to mind the scenario proposed by Wolstenholme (1996): true fetal mosaicism is typically associated with low levels of trisomy 8 in trophoblast cells (shortterm CVS culture), high levels in extraembryonic mesoderm (long-term CVS culture), and low levels in amniocytes and fetal blood cells.
Mosaic Trisomy 9 at Chorionic Villus Sampling
Saura et al. (1995) presented seven cases of trisomy 9, five of which gave a nonmosaic result, with the outcomes being abnormal in most. In the EUCROMIC study, of nine cases with trisomy 9 at either or both short- and long-term culture (including three with nonmosaic trisomy in one or both cultures), one proved to have fetal involvement (Hahnemann and Vejerslev, 1997). This single case had nonmosaic trisomy at both short- and long-term culture. Slater et al. (2000)report a case of trisomy 9 nonmosaic at CVS, but with level II mosaicism found at amniocentesis, with only 2 cells 47,XX,+9. At fetal blood sampling, all 85 cells analyzed were 46,XX. Molecular studies revealed upd(9)mat. A blood sample from the newborn infant had the karyotype 47,XX,+9[4]/46,XX[50]; upon review of the fetal blood material, 3 out of 102 cells were trisomic 9. Minor anomalies were noted in the child, who had been followed up to age 1 year. It is probable that this phenotype reflected a minor degree of residual trisomy in the child's soma.
Mosaic Trisomy 12 at Chorionic Villus Sampling
Hahnemann and Vejerslev (1997) and Sikkema-Raddatz et al. (1999) describe three cases, two of which involved a true fetal mosaicism. Of these latter, one fetus appeared grossly normal post-termination, and one infant was abnormal.
Mosaic Trisomy 13 at Chorionic Villus Sampling
A high level of trisomy 13 cells may well reflect significant mosaicism of the fetus. Ultrasonography, and amniocentesis with FISH analysis may clarify the picture. Mosaic trisomy 13 may present a very abnormal postnatal phenotype (Delatycki and Gardner, 1997). A difficulty arises in the case of very low–level (1% or so) mosaicism, in which case it is possible the child could be normal (Delatycki et al., 1998). In the EUCROMIC study, of 15 cases with trisomy 13 at either or both short- and long-term culture (including four with nonmosaic trisomy in one culture), 2 (14%) proved to have fetal involvement (Hahnemann and Vejerslev, 1997).
Mosaic Trisomy 14 at Chorionic Villus Sampling
Only three examples of 47,+14/46,N were recorded in the EUCROMIC study, none showing fetal trisomy (Hahnemann and Vejerslev, 1997). A theoretical risk exists for fetal UPD14 following “correction,” and two such cases associated with an initial CVS diagnosis are known. In the case of Morichon-Delvallez et al. (1994), the CVS was followed by a 46,XX karyotype at amniocentesis. The child proved to be normal apart from growth retardation, and maternal uniparental heterodisomy 14 was shown (Engel and Antonarakis, 2002). Ralph et al. (1999), diagnosing mosaic trisomy 14 at CVS, proceeded to follow-up amniocentesis, which also showed the mosaicism, and maternal uniparental isodisomy 14 was demonstrated. Fetal death in utero supervened; no morphological abnormality was identified.
Mosaic Trisomy 15 at Chorionic Villus Sampling
In a EUCROMIC study, 17 cases of trisomy 15 CPM were examined, in which direct and long-term cultures had been done (European collaborative research on mosaicism in CVS (EUCROMIC), 1999). In one a true fetal mosaicism was demonstrated. In about 30% the mosaicism was confined to trophoblast, and in about 25% to the villus mesoderm. In these circumstances, and if the level of trisomy is not high, the risk for abnormality, including due to UPD, appears to be small. A mitotic origin is probable. In the remaining nearly 50%, both trophoblast and villus mesoderm are involved, and here the risk for abnormality in the child is greater, especially if the trisomic load is high. These cases may well be of meiotic origin, thus setting the stage for possible “correction” and generation of UPD. Irrespective of these apparent distinctions, the recommendation was that amniocentesis be offered to all patients with a CVS diagnosis of mosaic trisomy 15, prudently to check for the possibility of UPD and true fetal mosaicism.
Mosaic Trisomy 16 at Chorionic Villus Sampling
Almost all CPM for trisomy 16 is due to a maternal meiosis I nondisjunction. A poor pregnancy outcome is very often observed (Benn, 1998; Hsu et al., 1998). In Benn's review, only 28% of cases of mosaic trisomy 16 at CVS were associated with a normal full-term pregnancy outcome; in Hsu et al., none was. Various malformations, intrauterine growth retardation, fetal death in utero, and fetal distress leading to emergency intervention have been observed. There has been debate about the relative roles of maternal UPD 16 (following “correction” of the trisomy), and of placental dysfunction due to placental trisomy 16, in bringing about these untoward effects. Given that the UPD cases are more severely affected than those with biparental inheritance, it is plausible that the UPD does, of itself, compromise fetal development, over and above any influence due to the placental insufficiency (Engel and Antonarakis, 2002; Yong et al., 2002). Yong et al. observe that knowledge of parental disomy status would not necessarily influence management of the pregnancy, and so they “do not yet advocate” prenatal testing for UPD upon the discovery of trisomy 16. DeLozier-Blanchet (2002) takes a less conservative view, and would offer UPD analysis; and it is true that, according to present understanding, an even more guarded prognosis would be given were UPD 16 to be proven. Additional to any effect of the UPD and placental insufficiency, a degree of fetal trisomy 16 mosaicism, even if this is very low, may be a further contributory factor. There may be what Benn calls “occult mosaicism”—a supposition of mosaic trisomy 16 in the child, albeit that cytogenetic analysis may have returned a normal result. Long-term data on surviving children are inadequate, and this needs remedying (Yong et al., 2003).
Mosaic Trisomy 18 at Chorionic Villus Sampling
In the EUCROMIC study, of 29 cases with trisomy 18 at either or both short- and long-term culture (including 8 with nonmosaic trisomy in one or both cultures), 4 (14%) proved to have fetal involvement (Hahnemann and Vejerslev, 1997). Harrison et al. (1993) studied placental karyotypes from pregnancies in which trisomy 18 had been diagnosed, either at pre- or postnatal diagnosis, and mosaicism was detected in 7 of 12, involving the cytotrophoblast. This supports the view that mosaic trisomy 18 at CVS may on occasion reflect a full trisomy of the fetus (and also leads to the conclusion that fetal survival may, in the context of this particular trisomy, be enhanced if there is a diploid placental fraction).
Mosaic Trisomy 20 at Chorionic Villus Sampling
In the EUCROMIC study, of 12 cases with trisomy 20 at either short-term, or at both short- and long-term culture (including four with nonmosaic trisomy in short-term culture), one (8%) proved to have fetal involvement (Hahnemann and Vejerslev, 1997). Steinberg Warren et al. (2001) described a child, normal apart from hypomelanosis of Ito, from a pregnancy with a nonmosaic trisomy 20 at CVS. As the skin sign indicates, the child was in fact mosaic, and this would probably have been revealed had amniocentesis been done (see Mosaic Trisomy 20 at Amniocentesis, below).
Mosaic Trisomy 21 at Chorionic Villus Sampling
Chromosome 21 naturally commands special attention. In the EUCROMIC study, of 22 cases with trisomy 21 at either or both short- and long-term culture (including 8 with non-mosaic trisomy in one culture), 9 (40%) proved to have fetal involvement (Hahnemann and Vejerslev, 1997). Beverstock et al. (1998) report a “near false-negative” finding of mosaic trisomy 21, in which trisomic cells were observed in only long-term CVS culture, and then, at follow-up amniocentesis, in only one culture. True mosaic trisomy was proven at fetal blood sampling and tissue culture post-abortion.
Mosaic Trisomy 22 at Chorionic Villus Sampling
Fetal defect is typically associated, but the degree may vary considerably. Wolstenholme et al. (2001a) described their own case of non-mosaic trisomy 22 diagnosed at direct and cultured CVS, with 47,XX,+22/46,XX mosaicism subsequently shown at amniocentesis (3/60 cells +22) and fetal skin biopsy (6/170 cells +22). Fairly subtle fetal dysmorphism was noted post-termination, and multiple tissue samplings showed mostly low but consistent trisomy mosaicism: 1% trisomic cells in skin, muscle, blood, kidney, 3% in lung, 5% in liver, and 21% in spinal cord. It is probable that neurological compromise would have transpired, quite likely of severe degree, had the child been born. Wolstenholme et al. reviewed 11 other cases of mosaic and nonmosaic trisomy 22, the mosaicisms mostly being of high percentages at CVS, and (in the six cases proceeding to amniocentesis) low percentages at amniocentesis. Of nine cases in which post-termination samplings were done, six showed mosaicism in at least some tissues (see also the case of De Pater et al., 1997, on p. 402, describing multiple placental samplings). In the three cases with 0% trisomy at fetal sampling, all had manifested severe IUGR. This may have been the consequence of functional insufficiency of the trisomic 22 placenta; there is also the point that occult fetal trisomy can never be excluded. Bryan et al. (2002) studied a child born of a pregnancy with a nonmosaic 47,XY,22 karyotype having been shown at CVS. There was IUGR, but the child apparently showed postnatal catch-up. He typed 46,XY on peripheral blood (with biparental disomy), and was phenotypically normal, except for hypospadias.
Detection at Amniotic Fluid Cell Culture
Considering the three major trisomies, Hsu et al. (1992) have determined that mosaicism for chromosomes 13, 18, and 21 very frequently predicts fetal abnormality, in half or more of cases. As for rare trisomies, Hsu et al. (1997) have undertaken a wide survey, based on the experiences of a number of American and Canadian laboratories and drawing on previous reports in the literature; the reader wishing full detail will need to refer to the original document. Some mosaic trisomies are associated with a high risk for phenotypic abnormality in the fetus or term infant, with figures of >60% for mosaic trisomies 2, 16 and 22, while trisomies 7, 8, and 17 are toward the lower end of the scale (<20%). Ultrasonography has a role in the assessment: most cases in which the mosaicism involves the fetus to a substantial degree will display morphologic/growth abnormality. Nevertheless, normal ultrasonography can by no means allow firm reassurance. Some mosaic states might cause structural defects too subtle to be discerned at fetal imaging, and yet be associated in the child with considerable, possibly severe functional neurological compromise. In chromosomes known to be subject to parent-of-origin imprinting, UPD needs also to be factored in to the assessment. Comments on individual trisomies follow.
Mosaic Trisomy 2 at Amniocentesis
In Hsu et al.'s (1997) survey, trisomy 2 conveyed the highest risk of any of the “rare trisomic” autosomes for an abnormal outcome, namely 90%, with a variable pattern of major defects. It is probable that mosaic trisomy 2 detected at amniocentesis would be in the same group as the high-level mosaic CVS case (see above). A trisomic line in the fetus/child may take some diligence to find. Sago et al. (1997) reported a case in which there was level III mosaicism with trisomy 2 cells present in 27% of amniocytes (and biparental disomy). The child was severely abnormal, and while blood and skin karyotyped as 46,XY, 4% of liver cells were 47, + 2.
Mosaic Trisomy 3 at Amniocentesis
Only two cases were identified in Hsu et al.'s (1997) review, in one of which the child had multiple malformations, with the mosaicism confirmed on skin fibroblast culture. The child in the other case was normal.
Mosaic Trisomy 4 at Amniocentesis
Zaslav et al. (2000) reviewed the three cases known at the time. In each, 47,+4 cells comprised the minor cell line (30%, 10%, and 8%, respectively). The first had an abnormal pregnancy outcome (dysmorphism and malformations), and mosaicism was shown on skin fibroblast culture. In the latter two cases, the infants were normal at 1 year follow-up, and foreskin and/or peripheral blood samples were nonmosaic 46,XY.
Mosaic Trisomy 5 at Amniocentesis
Hsu et al. (1997) recorded five cases. In one, a high level of trisomic cells (80%) was associated nevertheless with a phenotypically and karyotypically normal infant. In two cases, the child was abnormal, both showing the mosaicism on postnatal study.
Mosaic Trisomy 6 at Amniocentesis
Hsu et al. (1997) recorded three cases, each with the same low-level (6%) trisomy in amniocytes, and each with a normal outcome. But had amniocentesis been done in the CVS case of Miller et al. (2001) noted above, mosaic trisomy would probably have been seen, and hence, in this case, associated with phenotypic defect. In a case from Wallerstein et al. (2002), presenting for amniocentesis following the recognition of minor ultrasonographic signs, 5 out of 15 colonies showed trisomy 6, and the aborted fetus showed minor anomalies. No trisomic cells were observed on skin karyotyping, whereas all cells on placental analysis were 47,XX,+6.
Mosaic Trisomy 7 at Amniocentesis
Hsu et al. (1997) recorded eight cases, with fractions of trisomic cells ranging from 5% to 48%. Only one resulted in the birth of a phenotypically abnormal child, but low-level 47,XY,+7/46,XY mosaicism was confirmed in two phenotypically normal children on foreskin analysis. Warburton (2002) emphasizes that this relatively low risk assessment is the appropriate one to offer, and she notes also that UPD7, while unlikely, may be worth testing for. Mosaicism was also verified postnatally on skin fibroblast analysis in the child reported in Kivirikko et al. (2002), in whom fetal blood sampling and midtrimester ultrasonography had been normal; there was facial asymmetry and mild dysmorphism along with rather impressive hypomelanosis of Ito, while mental development was “considered to be within normal limits,” although no detailed assessment had been done. The fraction of trisomic colonies in the 47,XX,+7/46,XX case of Bilimoria and Rothenberg (2003) was rather high, at 41%, and in addition uniparental heterodisomy was shown in the 46,XX line; the pregnancy had come to attention because of an increased-risk interpretation from a maternal serum screen. On a neonatal blood sample, all cells were 46,XX, while all placental cells analyzed were trisomic. The child was small for dates and had some minor anomalies. These authors mention an anecdote of a trisomy 7 mosaic woman “graduating from college and getting married.”
Mosaic Trisomy 8 at Amniocentesis
Counseling is difficult, and advice must be cautious. An observation of trisomy 8 in amniocytes predicts a distinct probability, but by no means a certainty, of the clinical syndrome. It is not possible to put a good figure on the level of risk. Vice versa, a true fetal mosaicism may not necessarily be detected at amniocentesis (Wolstenholme, 1996). A finding of apparently normal morphology at fetal examination following termination in some 47,+8/46,N pregnancies is probably misleading, since the physical component of the clinical syndrome is relatively minor (Hsu et al., 1997). In the series of van Haelst et al. (2001)mentioned above, the two cases of trisomy 8 mosaicism detected at amniocentesis both turned out to be pseudomosaicism.
Mosaic Trisomy 9 at Amniocentesis
The risk is high (Saura et al., 1995). Hsu et al. (1997) recorded data on 25 cases, with pregnancy termination being done in 21. Abnormality was identified in most of the 21, and mosaicism confirmed in the seven having skin fibroblast studies. In the four pregnancies continuing, one abnormal child was born, with 47,+9/46,N mosaicism on blood karyotyping, the other three pregnancies resulting in apparently normal newborns. An overall figure of 56% applies for the risk that the fetus is abnormal. This high percentage figure is not surprising, and indeed it may well be an underestimate of the risk for functional abnormality in the child (intellect not being assessable in the newborn), considering the well-recorded phenotype of mosaic trisomy 9 in older individuals.
Mosaic Trisomy 11 at Amniocentesis
Hsu et al. (1997) recorded only two cases, both having low levels of mosaicism in amniocytes (3% and 5%), and both with a normal outcome. UPD 11 remains a theoretical consideration.
Mosaic Trisomy 12 at Amniocentesis
This is one of the more frequently described mosaicisms, and implies a high risk. Hsu et al. (1997) accumulated 23 cases, comprising 12 continuing pregnancies and 11 terminations. In most of those proceeding to fetal or neonatal fibroblast karyotyping, the mosaicism was subsequently confirmed, albeit that most of the fetuses appeared to be normal. It seems probable, however, that some subtle physical features, and possibly unsubtle neurological deficit, might have eventuated had these “normal” fetuses been born. The clinical range in the few recorded liveborn patients with true trisomy 12 mosaicism is very variable, from lethality in the newborn period, through to an otherwise normal man with Kartagener syndrome being investigated for infertility (DeLozier-Blanchet et al., 2000). Of the 12 continuing pregnancies in Hsu et al., the outcomes were abnormal in 5, and grossly normal newborns in 7. Three of these normal infants followed for 5 months to 5 years were all judged to be continuing to be normal. The proportion of trisomic cells at amniocentesis is not a very helpful guide in prognosis.
Mosaic Trisomy 13 at Amniocentesis
The risk for abnormality is very high. A collaboration of 23 American and Canadian laboratories provided data on the outcomes of 25 prenatal diagnoses of 47,+13/46 mosaicism (Wallerstein et al., 2000). Care was taken to exclude cases in which ascertainment had been biased by abnormal ultrasonography. In 21, the pregnancies were terminated. Various abnormalities were identified in 10 of these; the range of percentages of abnormal amniocytes was very wide, 6%–94%, average 58%. No defect was detectable in the remaining eleven aborted fetuses, although the assessment was limited to simple inspection. Four pregnancies proceeded to apparently normal live birth; the percentages of abnormal amniocytes in these were lower, ranging from 5% to 13%.
Mosaic Trisomy 14 at Amniocentesis
Hsu et al. (1997) recorded data on five cases. In the three choosing to continue the pregnancy to term, the infants appeared normal, and typed 46,N. In the two opting for termination, fetal abnormality was shown, in one case comprising hydrocephaly. A risk exists for UPD 14, over and above any defect due to the mosaic trisomy per se, and this should be checked (note that hydrocephalus is a feature of upd(14)mat). In another case of mosaic trisomy 14 detected at amniocentesis, upd(14)mat was shown in the normal cell line, and the pregnancy terminated (Sirchia et al., 1994).
Mosaic Trisomy 15 at Amniocentesis
Trisomy 15 is usually the consequence of a maternal meiosis I nondisjunction. Amniotic fluid mosaicism may well reflect a true mosaicism of he fetus. In Hsu et al. (1997), 6 of the 11 cases recorded had an abnormal outcome, the risk being greater when the trisomy level was higher (> 40%). Zaslav et al. (1998) review seven cases of low-level mosaic trisomy 15 detected at PND, in each the amniocentesis having been done for advanced maternal age. All seven chose to terminate, and a variety of defects were documented in most but not all. In their own case, the trisomic cell line in the initial amniocyte analysis was at a low level: 47,XX,+15[2]/46,XX[37]. Fetal tissues were also at low levels (lung 2%–5%, heart 8%–15%, skin 6%–10%, on metaphase and interphase analysis respectively), but the placenta showed 100% trisomy on metaphase analysis and 95% using FISH on interphase cells. These authors also document from the literature four cases of abnormal liveborns with trisomy 15 mosaicism. There is the additional question of UPD 15, the considerable phenotypic consequences of which may be superadded upon that of a trisomy 15 mosaicism.
Mosaic Trisomy 16 at Amniocentesis
In Hsu et al. (1997), 21 cases were documented, and abnormal outcomes were in the majority. Of 13 pregnancies continuing, 8 produced abnormal babies, and of 8 terminations, fetal defect was identified in all but 1, for a 71% abnormality rate overall. Benn (1998) records 29 cases of trisomy 16 mosaicism identified at amniocentesis. Most of the procedures had been done for the indication of advanced maternal age or because of an abnormal result on maternal serum screening. Only four cases had a normal outcome, in three of which the proportion of abnormal cells was low (1%–10%), but perplexingly in the remaining one, 92% of cells, and upon resampling, 73% were 47,+16. None of these four had follow-up cytogenetic studies. The overall risk is thus 86%, a very high figure. Hsu et al. (1998) report a rather similar experience with 11 cases, and there was an abnormal outcome in all the 5 pregnancies proceeding through to live birth. These authors comment that “mosaic trisomy 16 detected through amniocentesis is not a benign finding but associated with a high risk of abnormal outcome, most commonly intra-uterine growth retardation, congenital heart defect, developmental delay, and minor anomalies.” Uniparental disomy and placental dysfunction may make a contribution to this pathology, but fetal trisomy 16 mosaicism is the more severe factor (Engel and Antonarakis, 2002).
Mosaic Trisomy 17 at Amniocentesis
Hsu et al. (1997) record seven cases, with fractions of trisomic cells ranging from 5%–25% in six and 88% in the seventh. None displayed any fetal or neonatal abnormality. Genuardi et al. (1999) described three cases. In two, cordocentesis gave a normal karyotype, and in due course phenotypically and karyotypically normal children were born. In one, term placenta and amnion were 47,+17[1]/46,N[99], confirming confined placental mosaicism. One child had upd(17)mat, and this pregnancy was probably 47,+17 at conception due to maternal meiotic II error, with postzygotic “rescue.” In the third, the pregnancy was terminated, and fetal skin fibroblasts showed 47,+17 in 5%. Although the risk is apparently low, Hsu et al. do comment that trisomy 17 mosaicism has been recorded in abnormal liveborn infants, and the diagnosis “should not be taken lightly”; the third case of Genuardi et al. might well have been associated with abnormality, had the pregnancy continued.
Mosaic Trisomy 18 at Amniocentesis
The risk is very high. In the collaboration of Wallerstein et al. (2000), 31 prenatal diagnoses of trisomy 18 mosaicism were available for review. In just over half of these, the abortuses (induced termination or natural abortion) were abnormal. In 11, no defects were discerned at fetal examination. Just three pregnancies came to live birth, and these babies were apparently normal. The percentages of trisomic amniocytes in these three cases ranged from 2% to 20% (mean 9%), compared with 2%–95% (mean 37%) in those with abnormal outcome.
Mosaic Trisomy 19 at Amniocentesis
A single case is recorded, having a normal outcome at live birth (Hsu et al., 1997).
Mosaic Trisomy 20 at Amniocentesis
This is one of the more commonly observed mosaic aneuploidies. Trisomy 20 may exist in three forms: as confined placental mosaicism, as placental–fetal mosaicism with an apparently normal phenotype in the child that is subsequently born, or as a fetal mosaicism with phenotypic consequence (Hsu et al., 1991). There may be no dysmorphic features, or only some “soft” signs, or rarely an unambiguous facial dysmorphism. In certain fetal regions in which the trisomy may exist, in particular kidney and gut, the imbalance apparently has no discernible untoward effect, and in fact aneuploid cells may be cultured from urinary sediment. (Recognizing that amniotic fluid has a substantial contribution from fetal urine production, presumably some of the “amniotic fluid cells” from which the diagnosis of trisomy 20 had been made may have actually had origin from the fetal urinary tract.) In the collaboration of Wallerstein et al. (2000) comprising 152 diagnoses, 10 (7%) were recorded with an abnormal outcome (6 liveborns, 4 abortuses). There was correlation with the level of mosaicism: abnormality was observed in 20% of infants where there had been >50% trisomic cells at amniocentesis, and in 5% of those with <50%. Baty et al. (2001) reviewed 17 cases in which follow-up of the children extended beyond 1 year, of whom 12 (71%) had developed normally. The remaining five had various degrees of speech and motor delay. Fetal blood sampling is not helpful, as the trisomic cells rarely appear in blood.
Reish et al. (1998) offer the sobering example of a 15-month-old child with considerably delayed gross and fine motor skills and poor language acquisition, who had 54% trisomic cells from a skin biopsy (a normal karyotype on peripheral blood). In the pregnancy, amniocentesis had shown a 45% mosaicism, fetal ultrasonography was normal, and the parents had been “cautiously counseled.” To the contrary, Baty (2001) studied two cases with levels of trisomy of 83% and 57% at amniocentesis in one, and of 90% in the other, in which the outcomes were more fortunate. The two children, studied at ages 9 and 8 years, respectively, were of normal intelligence, and of essentially normal morphological appearance. Each did, however, display quite prominent hypomelanosis of Ito, presumably reflecting a fairly widespread distribution of a trisomic 20 lineage, at least in skin. A child followed up for 8¾ years by Steinberg Warren et al. (2001) had a similar phenotype, in this case following nonmosaic trisomy 20 at prenatal diagnosis (both CVS and amniocentesis). Two cases associated with UPD 20 are recorded, both having high levels of mosaicism in amniocytes, 85% and 98%, respectively (Salafsky et al., 2001; Velissariou et al., 2002). One of these children, 46,XX on blood, was developmentally delayed, and the other with 10% 47,XX,+20/46,XX mosaicism in blood (and 100% trisomy 20 in cells from urinary sediment) manifested a number of minor malformations as well as abnormal functional neurology at age 9 months; a contribution to the phenotypes in these children due to the UPD remains a moot point (see also p. 329).
Mosaic Trisomy 21 at Amniocentesis
The risk for Down syndrome is very high. The collaborative study of Wallerstein et al. (2000) accumulated 96 cases for review. Half had an observably abnormal outcome, with confirmatory cytogenetic study performed in a minority. Most of these were fetuses post-termina-tion with various abnormalities; six were liveborns, five of these having a clinical diagnosis of DS, and one an isolated heart defect. An apparently normal appearance (assessment limited to inspection in 39, autopsy in two) was recorded in 41 aborted fetuses. Among these, 20 were submitted to further cytogenetic analysis (repeat amniocentesis, fetal tissue, fetal blood, placenta), with eight showing 8%–90% trisomic cells, and 12 with 0%. Seven liveborns were normal, two being followed up beyond the newborn period; none had confirmatory karyotyping. The mean amniotic fluid proportion of trisomic cells was 17%, range 6%–31%, in these normal children. This compares with a mean of 35% in those with a demonstrably abnormal outcome. But even in the group with the lowest level of amniotic fluid trisomy, 3%–10%, half had an abnormal outcome. From the whole material, a risk for phenotypic abnormality of 50% should be seen as a minimum estimate, since subtler defects at fetal or neonatal assessment would have escaped notice, and a potential compromise of intellectual function of course was not assessable.
Mosaic Trisomy 22 at Amniocentesis
Hsu et al. (1997) determined a very high risk for abnormality for 47,+22/46, with 7 out of 11 outcomes being abnormal. Berghella et al. (1998) described a normal fetal blood result following trisomy 22 mosaicism diagnosis at amniocentesis, but fetal skin biopsy showed 47,+22/46, and structural abnormalities were subsequently identified in the aborted fetus. Four cases are noted in the review of Wolstenholme et al. (2001a), these all having followed an initial detection at CVS. Three out of the four showed some degree of normal/trisomy mosaicism at fetal samplings post-termination.
Mosaic Partial Trisomy at Amniocentesis
It is not feasible here to list recorded cases, and each must be judged on its merits. One specific example is worth noting, in that it may represent simply cultural artifact associated with a fragile site. This is mosaicism for a del(10)(q23). Zaslav et al. (2002) document a case of 46,XY,del(10)(q23)[9]/46,XY[45] detected at amniocentesis. The phenotypically normal child had the del(10q) in only 3/100 blood cells, this culture having been stressed by growth in a low-folate medium. We are aware of a handful of essentially similar cases, all involving 10q23, and none resulting in a documented abnormal child. The biology here is uncertain as amniotic fluid is normally cultured under conditions that suppress fragile site expression. Indeed, it is not clear that the known fragile site FRA10A at 10q23 is actually involved.
POLYPLOIDY
Triploidy
Close to 100% of the time, triploidy aborts spontaneously, but in some cases not until the pregnancy is well advanced. This being so, the offer of termination is appropriate when triploidy is diagnosed. Cassidy et al. (1977) described the emotional turmoil suffered by the family when a triploid infant, predicted to die immediately, survived for the extraordinary period of 5 months. Sarno et al. (1993) reported a unique case of complete placental/fetal discordance with triploidy on CVS and a normal diploid karyotype on amniocentesis and fetal blood sampling, with the birth of a normal baby; such a possibility warrants consideration where triploidy on CVS accompanies an ultrasonographically normal fetus. Nonmosaic triploidy typically shows ultrasonographic anomalies (Jauniaux et al., 1996).
Tetraploidy
Tetraploidy seen at PND, in the context of normal ultrasonography, is usually an in vitro cultural artifact, or possibly a vestige from the blastocystic stage of normally occurring trophoblastic tetraploidy (Benkhalifa et al., 1993). True tetraploidy is very rare, and Teyssier et al. (1997) record only 10 cases, 2 of which had been discovered at amniocentesis. Ultrasonographic demonstration of growth retardation and enlarged cerebral ventricles may be typical but rather nonspecific signs. While tetraploid/diploid mosaicism is almost always a cultural artifact, Edwards et al. (1994), having observed true normal/tetraploid mosaicism in two severely retarded individuals, nevertheless caution that a tetraploid cell line is not absolutely certain to be an innocuous finding. In a single such case at PND, Meiner et al. (1998) showed 92,XXYY/46,XY mosaicism on fetal blood sampling following the diagnosis of non-mosaic 92,XXYY at amniocentesis, in the setting of growth retardation discovered at ultrasonography and confirmed at subsequent fetal pathology study.
STRUCTURAL REARRANGEMENT
Structural rearrangements are seen in about 1 in 1000 cytogenetic prenatal diagnoses (Warburton, 1991). It is typically a matter of urgency to do parental chromosome studies, to distinguish between a familial or a de novo rearrangement in the fetus. If one parent is discovered to have the same apparently balanced rearrangement identified at PND, there is no firm evidence for an increased risk of fetal abnormality, and many would counsel to the effect of no discernibly increased risk. Possibly, a small, perhaps a per cent or so risk may apply due to a defect beyond the limits of routine cytogenetic resolution (see p. 94). Sex chromosome rearrangements require separate attention.
De Novo Apparently Balanced Structural Rearrangement
A major difficulty is posed by the rearrangement that at the level of cytogenetic analysis is “apparently balanced.” But even with the highest resolution banding, a submicroscopic abnormality (deletion or duplication, or gene disruption) may still be present (Wagstaff and Hemann, 1995; Kirchhoff et al., 2001). On postnatal observation, one can be wise after the event. If a child with a particular phenotype has a rearrangement involving a breakpoint known to be in the region of a Mendelian locus, or of other recorded rearrangements producing the similar phenotype, the conclusion could reasonably be drawn that the cytogenetic abnormality was the cause of that abnormal phenotype. For example, a child with a de novo inv(7)(p22q21.3) having a particular split hand/foot malformation would invite the inference of a causal link, given the similarity of limb defect with other 7q21.3–q22 rearrangements (Cobden et al., 1995). Sophisticated tools of the molecular cytogeneticist may reveal a hidden defect, such as an apparently balanced de novo 9q paracentric inversion in which Kleyman et al. (1997) could actually show a very small deletion. In a normal person, however, an apparently balanced rearrangement really can be supposed to be truly balanced.
Naturally, prenatal inference is less clear. Nevertheless, we should emphasize the observation that most pregnancies having PND of a de novo inversion or reciprocal translocation go on to produce a normal baby. Presumably, these normal cases reflect breakpoints in DNA that does not code for a gene or for a control element (or if a gene is disrupted, its haplostate is sufficient).
Warburton (1991) conducted a review of major laboratories in the United States and Canada over a 10-year period, and collected data based on more than a third of a million procedures. We make frequent reference to this work. A de novo translocation was identified in about 1 in 2000 amniocenteses, a Robertsonian translocation in about 1 in 9000, and an inversion in 1 in 10,000. She emphasizes that the outcome data are imperfect, given the lack of long-term follow-up and the questionable accuracy of phenotypic assessment in terminated pregnancies. Having made that point, she does say “there was no case in which a livebirth originally reported as normal was later classified as abnormal after longer fol-low-up. In fact, the opposite tended to occur: several cases described as having neonatal problems were later described as completely normal.” Small studies with follow-up into childhood have been undertaken (Gyejye et al., 2001), and suggest that the figures presently offered are in the vicinity of the truth, but a clear answer will require quite large numbers of children to be assessed. Given the long experience with prenatal diagnosis now accumulated, it is perhaps surprising that the data are as deficient as they are; or, if one considers the reality of what is involved in the logistics of long term follow-up, perhaps not.
De Novo Balanced Reciprocal Translocation
Simple Translocation
The starting point is that precedents (although not very many) are recorded for a de novo translocation having disrupted or compromised a locus, and therefore that the discovery of such a rearrangement at prenatal diagnosis could potentially herald an abnormal child. They are to be taken seriously. Equally, the balanced carrier state (all of which must have been de novo at some point in the near or distant past) is, of course, very familiar. Very many translocations are truly balanced, in terms of their functional genetic consequences. Thus, a normal child is very possible. In Warburton's study, serious malformations were identified in 6% of pregnancies with a de novo simple reciprocal translocation, either at elective termination or at live birth. This is some 3% above the background risk of around 3% for malformation and/or serious functional defect that applies to all pregnancies. Thus, we may draw the inference that in about 3% of these de novo translocations the chromosomal defect was causative. It seems reasonable to assume that a slightly higher figure, perhaps another per cent or so, should apply to the overall risk for not only major malformation, but also important functional deficit, which might not become apparent until after babyhood. Normal ultrasonography would be somewhat but not definitively reassuring.
Whole-arm Translocation. Very few de novo whole-arm translocations are recorded, “although the existing examples suggest an optimistic prognosis can be given” (Farrell and Fan, 1995). A whole-arm X-autosome translocation is mentioned below.
Complex Rearrangement. A de novo apparently balanced complex chromosome rearrangement (CCR) probably has a high risk for intellectual impairment and physical malformation, although prospective data are very meager. Ruiz et al. (1996) reviewed eight cases of CCR prenatal diagnosis, and three children had congenital defects, with neurodevelopmental deficiency in two of the five children assessed in this respect, for a total of 50% of children showing phenotypic defect. Madan et al. (1997) observe that, on postnatally studied cases, abnormal phenotypes are more associated with a greater number of breakpoints (mean 4.9) than are normal phenotypes (mean 3.6).
A 46,XX co-twin enabled a useful comparison in a pregnancy in which a de novo 46,XY,t(4;11;12;13) was identified in the other twin at 17-week amniocentesis (Peschka et al., 1999). Ultrasonography at 21 weeks showed normal fetal growth; but a repeat scan at 22 weeks indicated the onset of growth retardation, and “hectic abnormal movements” in the translocation twin. Rather subtle facial findings could be interpreted as abnormal in contrast to the 46,XX twin. Selective fetocide was done, and a normal girl and a fetus papyraceus were delivered at term.
Mosaicism for one of the Above De Novo Structural Rearrangements in Balanced State
Reciprocal Translocation Mosaicism
True mosaicism for a balanced reciprocal translocation, 46,rcp/46, is very rarely recognized (Fryns and Kleczkowska, 1986; Opheim et al., 1995; Leegte et al., 1998). The great majority of this type of mosaicism seen at prenatal diagnosis is level I or II, and is pseudomosaicism due to in vitro change. Some breakpoints (6p21, 13q14) are preferentially involved (Benn and Hsu, 1986). In terms of implications for fetal phenotype, it can usually be disregarded. True mosaicism for a reciprocal translocation has been reported at prenatal diagnosis, and Hsu et al. (1996) accumulated 11 examples showing one normal cell line and one with a balanced autosomal translocation. In no instance in which the pregnancy proceeded (nine of the 11) was phenotypic abnormality observed. Concerning a possible risk for unbalanced progeny in the next generation if the gonad were involved, each such case would need to be individually assessed; the parents would need to know to give their child access to the information in due course.
Robertsonian Translocation Mosaicism
In four cases in Hsu et al. (1996) of diagnosis at amniocentesis of mosaicism for a balanced heterologous translocation, 45,rob/46, the outcome was normal in all (the mosaicism confirmed postnatally in the two infants studied). The specific translocations were 13q14q, 13q22q and 14q21q.
Whole-Arm Translocation Mosaicism. The mother reported in Wang et al. (1998) with 46,XX,t(10q;16q)/46,XX mosaicism was normal (although her child abnormal; see p. 87). We know of one case of level III mosaicism for a whole-arm translocation at amniocentesis, 46,XY,t(1;5)(p10;q10)/46,XY, with 30% of cells in three separate cultures showing the translocation, and confirmed on a cord blood sample at delivery (10 cells out of 50 with the translocation); on follow-up at age 4 years the child was normal and healthy (D. Grimaldi and B. Richards, pers. comm., 2001).
Complex Rearrangement Mosaicism. The only known example of mosaicism with a CCR and a normal cell line detected prenatally is that described in Hastings et al. (1999b), and this case was associated with fetal abnormality.
Inversion Mosaicism. In four cases in Hsu et al. (1996) of diagnosis at amniocentesis of mosaicism for an inversion (pericentric or para-centric), 46,inv/46, the outcome was normal in all (all four were studied postnatally, with the mosaicism found in only one).
De Novo X or Y to Autosome Translocation
X-Autosome Translocation
In the case of a de novo apparently balanced X-autosome translocation, there are the additional possible complications of (a) gonadal dysgenesis if the breakpoint is within the critical regions of the X chromosome, (b) the unpredictability of the patterns of inactivation with the possibility of severe abnormality, and (c) a Mendelian disorder if the breakpoint iswithin a gene (see also Chapter 5). In one prospectively identified case in the series of Hsu et al. (1996), a normal male child was born following a prenatal diagnosis of 46,Y,t(X;9) (p21;q21)/46,XY. Hatchwell et al. (1996) provide the particular example of a severe phenotype associated with a whole-arm X-autosome translocation. On theoretical grounds, the risk may be about twice that for the simple autosomal translocation given above (Waters et al., 2001). Hatchwell et al. (1996) provide the particular example of a severe phenotype associated with a whole-arm X-autosome translocation.
On the specific issue of an Xp21 breakpoint, the question of Duchenne muscular dystrophy arises. Evans et al. (1993) actually showed normal dystrophin on a fetal muscle biopsy following detection at amniocentesis of an apparently balanced rcp(X;1) with the X breakpoint at p21, and so predicted the child would not have Duchenne/Becker muscular dystrophy; their prediction proved to be correct. In a case of de novo 46,X,t(X;9)(p21.3;q22) diagnosed at amniocentesis, Feldman et al. (1999) showed apparent integrity of the dystrophin locus on FISH. Methylation analysis indicated preferential inactivation of the normal X. On these two observations, the couple decided to continue the pregnancy; but fetal demise occurred at 34 weeks, probably due to chorioamnionitis following premature rupture of membranes at 33 weeks. No fetal defects were seen; dystrophin staining of muscle was normal.
Yq-Autosome Translocation
The important form of Yq-autosome rearrangement is the balanced reciprocal translocation, a 46-chromosome count, the breakpoint being in proximal Yq (the breakpoints usually given as q11, q11.2 or q12). Hsu (1994) reviewed 23 reports, in which the usual ascertainment was through infertility (oligospermia/azoospermia) in the adult, with a few being found incidentally and including one at prenatal diagnosis. Only three, including two from the early 1970s in which the detail of the rearrangement was less certain, were identified through a malformed child. It may be that such translocations should be regarded as conveying no greater risk for an abnormal intellectual phenotype than do reciprocal autosomal translocations. Fertility, however, may rather frequently be compromised (and see p. 113). In the particular case of a de novo translocation with Yqh material on the short arm of an acrocentric, this would be unlikely to be the basis of a phenotypic defect (and see p. 115).
De Novo Balanced Robertsonian Translocation
Heterologous Robertsonian Translocation
The additional risk for phenotypic defect is less than 1%, with the 95% confidence limits of this estimate encompassing 0%. That this risk figure is very small is not surprising, since the formation of the Robertsonian translocation classically does not disrupt unique-sequence DNA. Indeed, the figure might truly have been 0%, were it not for the possibilities of UPD and occult trisomy (see p. 128). Silverstein et al. (2002) reviewed their own and others' experience, accumulating data on 315 prenatal diagnoses, de novo and inherited, and, making the assumption of no different risk vis à vis UPD, pooled the two groups. Among these 315, two had UPD (of chromosomes 13 and 14 respectively), giving a point risk estimate of 0.65%, with 95% confidence limits 0.2%–2.3%. Given these data, it may be warranted to check for UPD, more especially in the setting of one of the imprintable chromosomes (14 or 15) being a component of the translocation. UPD 15 can be tested at PND using DNA methylation analysis at the 5′ SNRPN locus (Glenn et al., 2000). Amniocytes rather than chorionic villi may be the preferable tissue to test (Silverstein et al., 2002).
As for occult mosaic trisomy, this is a state that may have arisen from an initially trisomic 46,rob conception. The trisomic chromosome may then be discarded at a postzygotic “correction,” with the conceptus now cytogenetically balanced (45,rob), but with the possibility remaining of mosaicism with an occult, or very low–level trisomic cell line(45,rob/46,rob). While this cannot absolutely be excluded, the recorded experience to date indicates that this is an exceptional complication, at least at a level that might have evident clinical consequences. No such case was discovered in a de novo rob in the series of Berend et al. (2000a) (there was a single case due to a familial translocation that did have low-level (4%) trisomy 13 mosaicism, along with UPD 13).
Homologous Robertsonian Translocation (or Acrocentric Long Arm Isochromosome)
A chromosome comprising two long arms of the same acrocentric chromosome may be either an homologous Robertsonian translocation or an isochromosome, rob(13q13q)5 or i(13q). If the formation of an homologous rob has been through the fusion of the maternal and paternal homologs, which of course must have occurred as a post-fertilization event, then the rearrangement manifestly has to be a true Robertsonian translocation. Such a case is recognized by the demonstration of biparental inheritance, and a phenotypically normal child is the expectation, other things being equal (Abrams et al., 2001). Reproductive difficulty would, however, be anticipated; this would apply to the normal child with either a homologous Robertsonian translocation or an isochromosome (see Chapter 6).
The importance of UPD depends on the chromosome involved. In Berend et al.'s (2000a) Robertsonian series there were six identified with a homologous translocation, all de novo, and four of these had UPD, two upd(13)pat and two upd(14)pat. Barring isozygosity for a single gene mutation (see below), normal outcomes are to be expected following PND of a Robertsonian translocation (isochromosome) comprising a chromosome not subject to imprinting (chromosomes 13, 21, 22). This is actually recorded for the i(13q) UPD (Berend et al., 1999). No PND reports exist for i(21q) UPD or i(22q) UPD, but the postnatal state of normality in each of these is known (Engel and Antonarakis, 2002). Isodisomy for at least part of the chromosome will exist in the i(13q) UPD, i(21q) UPD and i(22q) UPD states, and this raises the question of a risk, not readily quantifiable but perhaps in the decimal point percentage range, for a Mendelian autosomal recessive disorder due to isozygosity, the parent being heterozygous for the mutation in question. For the imprintable chromosomes 14 and 15, however, the risk for clinical defect is absolute following PND of i(14q) UPD and i(15q) UPD, and the clinical syndromes of UPD 14 or UPD 15, maternal or paternal, would inevitably ensue (Berend et al., 2000a; McGowan et al., 2002).
De Novo Balanced Inversion (Pericentric or Paracentric)
(Note that a supposed “inversion” detected in this setting may actually be an unbalanced translocation). The risk from Warburton (1991) for phenotypic abnormality associated with a de novo inversion is 9.4%, which is 6%–7% over and above the background risk. The numerator is however small, and the 95% confidence limits span 2%–25%. Since, in theory, a two-breakpoint inversion should not imply a greater risk than the 2-break reciprocal translocation, the figure for this latter category as noted above, namely 3% (or a little above), might reasonably be seen as appropriate also for the inversion. In the special case of the X inversion, there may be gonadal insufficiency in the female (Dar et al., 1988; Dahoun, 1990).
De Novo Balanced Insertion
Only one case is recorded, to our knowledge, of a de novo apparently balanced autosomal interchromosomal insertion detected prenatally (Hashish et al., 1992). The child proved to be phenotypically normal. Van Hemel and Eussen (2000), in their review of nearly 90 families with an interchromosomal insertion, note that of the 9 probands with congenital anomalies having a balanced insertion, 7 were de novo and only 2 familial. It might reasonably be suggested that the risk for the interchromosomal insertion (three breakpoints) would be similar or possibly a little greater than the de novo apparently balanced reciprocal translocation (two breakpoints). Recalling the 3% risk figure associated with the latter, perhaps a per cent or so point above this is a fair figure to offer for the risk of “unspecified malformation and/or intellectual deficit.” Gonadal insufficiency may accompany the de novo intrachromosomal ins(X) (Grass et al., 1981).
De Novo Unbalanced Structural Rearrangement
Unbalanced Rearrangement, Modal Number 46 or 45
Autosomal. For any de novo autosomal structural rearrangement in which cytogenetic imbalance can be demonstrated, serious phenotypic abnormality is highly likely. Often, it is not possible readily to identify the precise origin of a duplicated segment, which means that precise prediction of phenotype is not possible. Many cases, indeed most, are unlikely to be exactly the same as those in the literature or on the databases, and the counselor will need to make an informed evaluation. Ultrasonography may clarify the question if abnormalities are seen, but an apparently normal sonogram does not guarantee that the child would be normal (Al-Kouatly et al., 2002).
In the mosaic state, the risk may be high if pseudomosaicism is judged to be unlikely. Hsu et al. (1992) record 34 cases with at least one cell line having an unbalanced rearrangement (thus, presumed to be a true mosaicism). In follow-up studies, phenotypic abnormality was noted in about 50% and cytogenetic confirmation obtained in 65%. Each rearrangement needs to be considered on its merits. The dilemma of deciding how best to advise couples is illustrated in Cotter et al. (1998). They describe the karyotype 46,XX,der(4)t(4;5)(q34;q12)/46,XX detected at amniocentesis, imparting, in the abnormal cell line, trisomy for most of 5q. This was confirmed on two subsequent amniocenteses, with an average overall of 17% of amniocytes abnormal, but with a 46,XX result on fetal blood sampling, and normal ultrasonography. The parents were advised that “few data were available” to determine risk; they made a decision to continue the pregnancy. In the event, the child appeared normal at birth and at 2-year follow-up; 100 cells at cord blood karyotyping were normal. Cotter et al. rightly call for others' experience in similar cases to be published.
X-Autosomal. Prediction with respect to the X-autosome translocation is precarious. The degree to which selective inactivation may occur, and its extent in the fetus, is not knowable. Although the pattern of inactivation may lessen the effect, and indeed convert an invariably lethal imbalance to a survivable state, a significant defect remains very probable (Kulharya et al., 1995; Garcia-Heras et al., 1997; Orellana et al., 2001). Had the child with an unbalanced der(X)t(Xp;22q) described on p. 111 (Fig. 5-10) been identified at amniocentesis, and with the DiGeorge critical region intact and no inactivation on the 22q segment, a prediction of typical Turner syndrome might have been reasonable.
In the event, this child proved to have a significant mental handicap. Contrary examples in which a prediction of major abnormality would have been mistaken are rare, such as the cases outlined on p. 103.
Unbalanced Rearrangement, Modal Number 47—Extra Structurally Abnormal Chromosome
Extra structurally abnormal chromosomes are a heterogeneous group. They have been described variously as marker, supernumerary, accessory, and B-chromosomes (Hook and Cross, 1987a). Some are quite harmless, and associated with phenotypic normality (the B-chromosome), and others are not. ESACs are encountered in about 1 in 1000 prenatal diagnoses, frequently in the mosaic state with a normal cell line. Upon the discovery of an ESAC at prenatal diagnosis, an urgent parental chromosome analysis is required. The majority will prove to be de novo. The following questions should be asked: From which chromosome is it derived, and does it comprise euchromatin or heterochromatin? Is it a recognized type of ESAC, for which precedents are recorded? Precise characterization is necessary, and this requires the use of special stains. On FISH, about 80% are shown to derive from one of the acrocentric chromosomes, most commonly no. 15 or no. 22, and often involving only the pericentromeric region and/or the satellites (Crolla et al., 1998). If available, the technique of spectral karyotyping offers the possibility of rapid identification of the origin and nature of some of the ESACs (Haddad et al., 1998; Ning et al., 1999; Yaron et al., 2003). Gonosomal ESACs are discussed separately in the section on sex chromosomes below.
Familial Extra Structurally Abnormal Chromosome
Interpretation in the case of a familial ESAC is usually straightforward. If one parent is also 47,+ESAC and phenotypically normal, it can be assumed no discernibly increased risk for fetal abnormality exists (Brøndum-Nielsen and Mikkelsen, 1995). Hastings et al. (1999a) surveyed a 10-year experience in London, and report six familial ESACs, which included three that were 14 or 22 derived, an idic(15), a der(6) and a mosaic der (16). The outcomes in the five proceeding to live birth were all normal at follow-up from 5 months through 5 years. When the parent has the ESAC in mosaic state, prediction for the fetus is more difficult: the chromosome could be potentially harmful, but the parent might have been protected by a particular tissue distribution. The mosaic der(16) in Hastings et al. comprised centromeric chromatin: mother and child, both mosaic, were normal. Each case will need to be judged on its merits. The idic(22) presents an exception, since a parent can be normal and the child abnormal (Crolla et al., 1997). If the ESAC is revealed as being a small derivative chromosome from 3:1 malsegregation, one parent being a balanced translocation carrier (Stamberg and Thomas, 1986), serious phenotypic abnormality is practically certain.
De Novo Extra Structurally Abnormal Chromosome
De novo ESACs have been described for most chromosomes (Hastings et al., 1999a). The mode of ascertainment may suggest a category of risk: those in which fetal ultrasonographic anomaly has been detected would enter a higher-risk group, as might, intuitively, those discovered through an increased-risk finding at maternal serum screening. The risk for abnormality is low in the very small derivatives of acrocentric chromosomes which stain negatively for euchromatin, and which may be satellited. If, however, a der(15) contains the segment of proximal 15q that includes the Prader-Willi/Angelman region, the risk is high (see below). Mosaicism appears not to alter the risk for abnormality.
With a reasonable level of cytogenetic characterization of ESACs and ultrasound examination it is possible to categorize most fetuses as being either at high risk of abnormality, or at a relatively low risk (less or much less than 5%). In principle, those comprising heterochromatin convey a low risk, while a euchromatic ESAC may imply a high risk for phenotypic abnormality. Published series of liveborn children with ESACs are mostly biased by ascertainment in favor of phenotypic abnormality. Series of prenatally diagnosed fetuses are deficient in that there is usually only a short-term follow-up of liveborn children, while pathological assessments following termination can only show major structural malformations (Warburton, 1991).
Brøndum-Nielsen and Mikkelsen (1995) report a 10-year experience in Glostrup during which nine de novo ESACs were identified. In seven cases, termination of pregnancy was chosen, with some of these showing defects at pathological examination, and in the two pregnancies continuing, one infant with a minute acrocentric-derived ESAC was normal at birth, while one with a ring-like 17 was “slightly retarded” at age 2 years. In the similar survey of Hastings et al. (1999a), data were presented on 31 prenatally diagnosed ESACs, of which 21 were de novo. In 10 of these 21 proceeding to FISH analysis, 6 being mosaic, 5 were shown to be 15 derived and 3 were 14 or 22 derived; the remaining 2 included a r(8) and a der(16). Of the six in which the pregnancies continued, only the r(8) child was physically and developmentally abnormal.
Specific Well-Characterized De Novo Extra Structurally Abnormal Chromosomes
Isodicentric 15. About half of all ESACs are an idic(15) (also referred to as pseudodicentric 15, or inverted duplication 15). These are typically dicentric and bisatellited, although one of the centromeres may be suppressed. The smallest ones (smaller than chromosome 21q) appear to be harmless, but larger ones result in the “idic(15) syndrome,” characterized by mental defect and autistic features. The boundary between smaller and larger may be 15q12. The use of D15S10 or SNRPN FISH probes, which recognize sequences in 15q12–q13, enables distinction of harmless and pathogenic chromosomes (Eggermann et al., 2002). Rare idic(15)s have been associated with UPD 15, and it may be warranted to check for this possibility (Hastings et al., 1999a).
Isodicentric 22. The bisatellited idic(22) typically, but not invariably, causes cat-eye syndrome. If the idic(22) lacks proximal 22q euchromatin, normality is very probable, whereas those containing euchromatin can lead to a phenotype anywhere between full cat-eye syndrome and normality (Crolla et al., 1997).
Autosomal Isochromosomes
The mosaic state is usual for a supernumerary isochromosome, and thus the discovery of 47,+i/46,N is always a serious concern. Such a karyotype raises the prospect of an effective mosaic tetrasomy for the chromosomal arm concerned. A 46-chromosome karyotype in which one homolog is replaced by an isochromosome typically implies a trisomy for one arm of that chromosome, and monosomy for the other.
47,+i(5p). Sijmons et al. (1993) assessed a dysmorphic and neurologically compromised child with a 5p isochromosome in 3/31 lymphocytes and 12/14 skin fibroblasts, and yet upon retrospective checking, only one of 217 cells from a stored short-term CVS culture was 47,XY,+i(5p). We contrast this unfortunate experience with ours of seven cases of i(5p) mosaicism identified at CVS, six of which went on to follow-up amniocentesis (Clement Wilson et al., 2002). Three children were followed up to 2½, 3¼, and 4 years, and their normality was quite apparent. In one of these children, a circumscribed area of the placenta following delivery karyotyped 47,+i(5p), adjacent parts karyotyped 47,+i(5p)/46,N, and most of the placenta (and the child himself) had a normal karyotype. The CVS sampling had presumably needled this small region of confined placental i(5p) mosaicism. One pregnancy tested 100% i(5p) at CVS, and the parents chose termination; no i(5p) cells were detected from fetal skin culture. In another with a 65% load at CVS, a follow-up amniocentesis showed 45% of cells with the isochromosome, and post-termination tissues showed 15%–30%. From the foregoing, we may conclude that a CVS diagnosis with a normal follow-up amniocentesis and with normal ultrasonography suggests, but cannot confirm, a normal child. As for the primary detection of i(5p) mosaicism at amniocentesis, only three cases are recorded, all three having an abnormal outcome (Reddy and Huang, 2003).
47,+i(8p). López-Pajares et al. (2003) review the small number of reported cases. Two examples are given of discordance between amniocentesis (normal) and postnatal blood (tetrasomy 8p), a most unusual pattern, shared with the i(9p) following.
47,+i(9p). The clinical picture, and the subtleties of different breakpoints, are reviewed in Dhandha et al. (2002). Isochromosome mosaicism can be the basis of a false-negative test result at prenatal diagnosis. Thus, Eggermann et al. (1998) reported an abnormal baby born to a 39-year-old mother, in whom amniocentesis at 14 weeks gestation had returned a normal karyotype. On blood analysis, the child had an i(9p) in 32% of cells. From one skin biopsy, 50 cells had a normal karyotype, but on a second biopsy, 5 out of 8 cells showed the i(9p) chromosome. The particular attribute of the i(9p) is for blood, but not skin, to show the abnormality (in contradistinction, for example, to the i(12p) noted below), and this may provide the explanation for its non-detection at amniocentesis. Pertile et al. (1996) support this interpretation, in their follow-up of a (nonmosaic) CVS diagnosis of idic(9)(q13). An extensive search at amniocentesis revealed a single abnormal colony, which might well otherwise have been missed. Finally, fetal blood sampling showed the idic(9) in 8% of cells.
47,+i(10p). A single case is on record, the diagnosis having been made following the recognition of fetal defects on ultrasonography (Wu et al., 2003).
47,+i(12p). The 12p isochromosome is the basis of the Pallister-Killian syndrome. The fractions of abnormal cells detected at PND can vary greatly. Bernert et al. (1992) showed in one example 100% of short-term CVS cells and 10% of amniotic fluid cells having the 47,+i(12p) karyotype; the pregnancy was terminated. Horn et al. (1995) reported a pregnancy in which CVS gave a 46,XY result on direct (17 cells) and cultured (8 cells) analysis (and 28 further cells on a retrospective study), and the abnormal newborn baby was 46,XY on a peripheral blood study (100 cells counted); at 18 months, a clinical diagnosis of Pallister-Killian syndrome was made, and the karyotype on skin fibroblast culture was 47,XY,+i(12p)/ 46,XY, with 85% of cells having the isochromosome. (Had it been an amniocentesis rather than CVS that had been done, abnormal cells might well have been seen.)
47,+i(18p). Over 50 cases of 47,+i(18p) are recorded (Kotzot et al., 1996). Boyle et al. (2001) emphasize the plausibility of a premeiotic origin, and the caution therefore that gonadal mosaicism may exist in a parent, as they illustrate in their report of affected half-sisters.
46,i(18q). The karyotype produces a combination of monosomy 18p and trisomy 18q. Chen et al. (1998) record that many 18q isochromosomes diagnosed prenatally are associated with very severe malformation, such as holoprosencephaly and cloacal dysgenesis. Levy-Mozziconacci et al. (1996) describe a case presenting at 22 weeks gestation with abnormal ultrasonography, and although the direct CVS was 46,XX in all cells, amniocentesis and fetal blood sampling showed the isochromosome (an isodicentric, in this instance) in all cells; an example of complete CVS-amniocentesis discordance.
46,i(20q). An i(20q) identified at amniocentesis in mosaic form appears typically to be a benign finding, a rather surprising conclusion. It may be an unusual sort of mosaicism in being confined to amniocytes, and there are numerous examples of the subsequently born normal baby having a normal karyotype on peripheral blood (Chen, 2003). The inconsistent pattern of defects in the few cases with malformation may reflect coincidence, although a firm statement to this effect cannot presently be made.
46,i(21q). Although this rearrangement is sometimes called a Robertsonian translocation, in fact it is typically an isochromosome (Shaffer et al., 1991). The phenotype is that of typical Down syndrome. Gilardi et al. (2002) report a case in which the isochromosome probably arose postzygotically in an early cell destined to form the lineage of the inner cell mass and the extraembryonic mesoderm, such that a direct CVS gave a nonmosaic 46,XX result, while long-term CVS and post-termination fetal studies showed nonmosaic 46,XX,i(21q), and a similar story comes frim Brisset et al. (2003). The i(21q) can also exist in a 47-chromosome karyotype. Nagarsheth and Mootabar (1997) showed a 47,XY,+i(21q)[6]/46,XY[19] karyotype at amniocentesis; the parents elected to continue the pregnancy, and the abnormal child had only 1 out of 120 peripheral blood lymphocytes with the i(21q), the other 119 being normal. These authors suggest that some previously reported cases of supposed i(12p) mosaicism may have been, in fact, i(21q).
De Novo Supernumerary Ring Chromosomes
Supernumerary small distamycin A/DAPI6 negative ring chromosomes imply a high risk of phenotypic abnormality. They originate from a variety of chromosomes and contain euchromatin. Certain of these, in which only one arm of the chromosome is represented in the ring, are specifically recorded in association with phenotypic abnormality: r(1p), r(5p), r(7q), r(8q), r(9p), r(10p), r(20p), r(20q) (Anderlid et al., 2001). The r(8) with an abnormal outcome in Hastings et al. (1999a) is mentioned above. Uniparental disomy may complicate the picture: James et al. (1995) and Anderlid et al. (2001) report supernumerary rings, from chromosomes 6 and 9, associated with UPD 6 and UPD 9 respectively. Mosaicism for an apparently nondeleted ring chromosome can present a difficulty in interpretation. For example, Flejter et al. (1996) document the variable clinical picture in (postnatally diagnosed) 46,r(19)/46,N, which seemed not to correlate with the fraction of cells showing the ring (and see p. 181).
Chromosomal Breakage Resembling the ICF Syndrome. The chromosomes at CVS may on occasion somewhat resemble the “starburst” appearance in the ICF syndrome (p. 307, Fig. 19-4). Ehrlich et al. (2001) noted that “anecdotal observations of these types of pericentromeric chromosome 1 and 16 anomalies in normal CVS metaphases are common,” and concluded that “ICF-like chromosomal abnormalities are part of the normal spectrum for CVS chromosomes and need not indicate any clinical condition,” a conclusion supported by subsequent study (Tsien et al., 2002).
SEX CHROMOSOME ABNORMALITIES
Full Aneuploidy
A sex chromosome abnormality is not an uncommon discovery at prenatal diagnosis, with an overall incidence of 1 in 250–300 (Linden et al., 2002). The main conditions are XXY, XXX, XYY and 45,X. Two of these (XXY and 45,X) may be firmly predicted in terms of an abnormality of development of the reproductive system. Children with Klinefelter and 45,X Turner syndrome will with near-certainty be infertile. For those couples deciding to continue a pregnancy, Robinson et al. (1986) offer a useful commentary. Parents of children predicted to be infertile might feel a sense of loss—a “sadness and regret about their child's anticipated loss and about their own loss of grandchildren” and “concern about their children's wholeness and, by extension, their own.” Parents may take some comfort from knowing that infertility is by no means an uncommon problem in the general population, and further comfort from the advice that recent advances in artificial reproductive technology may now enable the infertility to be overcome in some individuals.
The picture for intellectual and psychological functioning is less predictable. Earlier adult studies defining a strong association with mental deficiency and psychological disturbance were contaminated by ascertainment bias (and counselors' personal experience may have been more with those children whose problems were sufficiently severe that they had come to medical attention). Children identified in newborn populations screened for cytogenetic abnormalities and subsequently followed up constitute a group unbiased in their ascertainment, although perhaps subject to other but less important biases (Puck, 1981). Data from the study of such children in several American and European cities, followed from infancy through childhood, adolescence, and young adulthood have now given a reasonably clear picture of the natural history of the more common sex chromosome aneuploidies (Linden et al., 2002).
In general, the IQ averages 10–15 points below that of the siblings. Hook's (1979) early proposition has held up: some sex chromosome aneuploidies influence brain function in such a way that the development of intellectual capacity, emotional maturity, and speech and language skills are affected to some extent; but none of these effects necessarily occurs, none is specific to sex chromosome aneuploidy, and some may be amenable to corrective intervention. There is considerable overlap with the XX and XY population! Ratcliffe (1999) and Bender et al. (2001) provide long-term follow up data, well into adulthood. Bender et al. followed 8 45,X, 10 47,XXX and 11 47,XXY individuals through to an age range of 26–36 years, using siblings as controls, and noted the IQs of the aneuploid groups to be considerably less compared with the sibs. Nevertheless, the variation is wide, and these authors emphasize the point that “sex chromosome aneuploidy does not exert its influence in a vacuum, but rather interacts with the host of other genetic and environmental influences that collectively guide human development.” Children with sex chromosome aneuploidies seem more susceptible to either the good or the bad effects of a stable or of a dysfunctional family setting than do their 46,XX and 46,XY siblings (Stewart et al., 1990; Bender et al., 1995). Children identified at prenatal diagnosis, a group biased towards higher socio-economic status, may do better academically and socially than the cohorts followed from birth, although it was nevertheless true in the study of Linden and Bender (2002) that these children had “a strong risk for developmental problems, particularly for learning disabilities [although] these problems were not often severe.”
If a couple decides to continue the pregnancy, what should they say to others? Should the family know, should they tell friends, and should school personnel be aware? And when should the child learn about their chromosomal condition? Linden et al. (2002) have considered these questions, and in general make a case for openness within the family, but see no need, indeed potential disadvantage, for those outside to be told.
We next outline the predicted outlook for the more commonly encountered sex chromosome aneuploidies. Attention is paid mostly to gonadal function and to intellectual and social development.
XXY (Klinefelter Syndrome)
Almost certainly, the child becomes an infertile adult, and while penile size is usually normal the testes will be small. Androgen deficiency can be managed by replacement therapy with testosterone. It may be that treatment induces a more masculine body habitus, improved self-esteem, vitality, ability to concentrate, and sexual interest (Nielsen, 1990; Winter, 1990). Gynecomastia may be present, transiently, in 50%; if it persists, it can be treated surgically.
IQ is diminished by some 10–15 points, with a particular impairment in language expression being common. Learning difficulty at school is to be expected. Of 13 XXY boys studied by Walzer et al. (1990), 11 had persistent reading and spelling problems. Specific characteristics included a lowered level of motor activity, a pliant disposition, and a cautious approach to new situations; thus, in the classroom setting, they are perceived as “low-key children, well liked by their teachers, and presenting few behavioral management problems.” Speculatively, the neural substrate of this passivity may reside in an underdevelopment of the amygdala (Patwardhan et al., 2002). Bender et al. (1993) note that a deficit in verbal fluency and reading is “the most homogeneous and consistent cognitive impairment found in any sex chromosome abnormality group,” and this may reflect a specific dysfunction of the left cerebral hemisphere.
Six Danish XXY boys were followed from birth to age 15–19 by Nielsen and Wohlert (1991), and all but one needed remedial teaching. Their career plans were carpenter, draughtsman, gardener, unskilled laborer, mechanic, and undecided. Stewart et al. (1990) comment that “XXY boys are unlikely to reach a level of personal and social development that is consistent with their family background.” Ratcliffe (1999) commented on a rate of psychiatric referral being above that of male controls (26% vs. 9%), with the neurotic score (not the antisocial score) being higher. (She also notes anecdotal mention of men from a Klinefelter clinic with professions including physician, engineer, minister, and accountant.) In a summary of psychosocial adaptation from several studies, recurring adjectives to describe the XXY personality were shy, immature, restrained, reserved. In the Denver study, 11 young adults with XXY “appeared to have met the demands of early adulthood with fair success, although slightly less well than did their siblings”; they appeared to have a diminished insight into their own psychology (Bender et al., 1999). Their mean IQ of 91 compared with 109 in normal male sibling controls. We have noted above the ameliorative effect of growing up in a stable and supportive family.
XXX
Physical development of the XXX female is generally unremarkable, although there is a tendency toward tallness. Gross and fine motor skills are likely to be somewhat impaired, and children are awkward and poorly coordinated. Pubertal development and fertility appear, for the most part, uncompromised. The major concerns relate to intelligence and language. Full scale and verbal IQ is reduced by some 10–20 points. Language comprehension and use of speech are impaired in over half the cases. Learning difficulty is likely and many will benefit from additional remedial teaching, but few require education outside the mainstream. In one small study of 11 girls, 9 needed special education intervention, and one was placed in a class for retarded children (Bender et al., 1993).
Harmon et al. (1998) and Bender et al. (1999) reported a longer follow-up in these young women, into adolescence and young adulthood, and documented difficult adaptation to the stresses of life. On a measure of social adjustment (in work, leisure, family, marital, parental), the XXX women scored significantly less well than their sisters. Their mean IQ was 82 (cf. sisters, 103). However, Ratcliffe (1999) described most XXX young women in the Edinburgh survey as “physically attractive, and displaying a common sense attitude that counterbalanced their low educational achievements” (and relieved to be free of the pressure they had felt while at school). The observations in the similar study of Rovet et al. (1995) were more promising, although, as Harmon et al. point out, this was a group from a higher socioeconomic stratum, and presumably both genetic and environmental factors would have been more favorable. An XXX girl who might otherwise have had an IQ of 130 can yet do well in spite of a reduction to 110; to the contrary, a drop from 90 to 70 would be a considerable handicap. Many counselors will know from their own experience how variable can be the phenotype.
XYY
The multicenter prospective study documented in Evans et al. (1990) reviewed progress in 39 boys and young men. The particular physical attribute of the XYY male is increased stature. Sexual activity is normal, and fertility is apparently uncompromised. Motor proficiency may be impaired. While the IQ is in the normal range, it is usually lower than those of sibs or controls, and about half of XYY boys have a mild learning difficulty. It may be that the aneuploidy causes a minor and subtle impairment of neurologic maturation, leading to some features of minimal brain dysfunction (Theilgaard, 1986). The vignettes from the series of Ratcliffe et al. (1990) of 10 Scottish subjects who have left school give an idea of what XYY young men are capable of: one runs a market stall, two are chefs, and the others are a private in the army, a waiter, a supermarket assistant, a video shop assistant, a technician, a laborer, and one training as a painter and decorator. In a cohort of children aged 8–16 selected for the XYY karyotype having been diagnosed prenatally, and of higher socioeconomic status, a considerable range in academic ability was observed, with most coping satisfactorily, and IQs ranging from 100 to 147 (Linden and Bender, 2002).
Perhaps the major concern is in psychosocial adaptation. These boys can have a low frustration tolerance, and some are prone to temper tantrums in childhood progressing to aggressive behavior in teenage, and may need help to learn to cope with this. Antisocial behavior is more common (Ratcliffe, 1999). The functioning of the family may be as much an ingredient as the karyotype in psychosocial development. Fryns et al. (1995) identified 50 XYY males among 98,725 patients referred for chromosomal analysis, and they note that this fraction of 50/98,725, approximately 1/2000, is very close to the newborn incidence, and thus drew a conclusion that the XYY phenotype differs little from the norm. They do, however, acknowledge a high (86%) risk for psychosocial pathology in those XYY males with concomitant borderline intelligence or frank mental deficiency. In Ratcliffe's (1999) follow-up report into adulthood, some disconcerting data are noted, not incongruent with the conclusions of Fryns et al. Psychiatric referrals were fivefold compared with male controls (47% cf. 9%), and the rate of criminal conviction fourfold, the mean IQ of those convicted being lower than those who were not (although most offences were minor, and against property rather than persons).
45,X (Turner Syndrome)
Unlike the foregoing aneuploidies, monosomy X has a very high in utero lethality, peaking at around 12 to 15 weeks gestation. Spontaneous abortion follows amniocentesis-detected 45,X in three-quarters of cases (Hook, 1983). But some survive pregnancy and are born as infants with Turner syndrome. Robinson et al. (1990) note that “variability among 45,X girls is considerable; and precise predictions about any child's prognosis are not possible.” They also emphasized that “a supportive environment that provides stimulation and encouragement is of considerable importance.” These traits comprise the core phenotype:
1. Stature will be short. In a study of adult Danish women with Turner syndrome, never having had growth hormone therapy, the average height (with standard deviation) was 147 cm ± 7 cm (4 feet 10 inches ± 2½ inches) (Gravholt and Naeraa, 1997), which may be a little taller than in some other populations. A useful increment can be achieved with growth hormone treatment.
2. Gonadal failure with infertility is almost certain (Lippe, 1991). Classically, a spontaneous onset of puberty, with breast development and onset of menses, has been regarded as being very infrequent. Latterly, Pasquino et al. (1997) proposed that the fraction who enter a spontaneous puberty may be as high as 9%, and they suggest that earlier figures may have been biased downwards by a policy, previously, of not karyotyping short girls who had had an onset of menstruation. Childbearing via ovum donation may be successful in some cases (Chapter 12). Pavlidis et al. (1995) reviewed sexual functioning in women with Turner syndrome, and suggest strategies to avoid possible difficulties.
3. The IQ is reduced compared to siblings. At long-term follow-up in the Denver cohort (Bender et al., 1999), nine young women with 45,X had a mean lower IQ (85) compared with normal female sib-ling controls (104). Their educational achievements were, however, better than those of the XXX women from the same study: eight were high school graduates, and five had college degrees. In one notable case, Reiss et al. (1993) report monozygous twins, one nonmosaic 45,X and the other 46,XX, the former's performance IQ being 18 points less than her sister, but the verbal IQs practically the same. Psychological assessment indicates a particular vulnerability in social adaptation (Bender et al., 1999). Reiss et al. (1993) review aspects of the cognitive–behavioral phenotype, and correlate the specific feature of difficulty with visual–spatial appreciation with a lesser volume of the right parietal cerebral cortex. Romans et al. (1998) confirmed and extended this appraisal in a study of 99 subjects with Turner syndrome, in whom they identified diminished abilities on measures of spatial and perceptual skills, visual–motor integration, recognition of facial expressions associated with a particular affect, visual memory, attention, and executive function (the ability to plan, organize, monitor, and execute multistep problem-solving processes). These traits are not improved by taking estrogen (Ross et al., 2002).
4. Certain physical defects are associated, of which the major are neck webbing and coarctation (narrowing) of the aorta.
5. Morbidity in adult life is increased (Gravholt, 2001; Swerdlow et al., 2001). Certain common diseases are more frequently seen: obesity, both insulin-dependant and insulin-resistant diabetes, hypothyroidism, heart disease, hypertension, stroke, and liver cirrhosis. Weakness of the bones (osteoporosis) implies a risk for fracture. There may be a place for ongoing hormone replacement therapy.
There is a possibility that Y-chromosome material may be present even if the karyotype is apparently nonmosaic 45,X. Huang et al. (2002)reviewed 74 cases of 45,X diagnosed prenatally, most having been ascertained via, or discovered with, abnormal fetal ultrasonography. Of six with normal ultrasonography, three showed a male genital phenotype. The explanations, upon more detailed analysis, were as follows: in one, a segment of Yp was translocated to a no. 14, shown on FISH with an SRY probe, and in the other two, there was low-level mosaicism for an idic(Y) marker. Normal male children were born. Some women with Turner syndrome who are 45,X on karyotyping may actually show Y sequences on molecular study, and these women do have a greater risk for gonadoblastoma (Mendes et al., 1999).
Sex Chromosome Polysomy
This category is to be seen as quite separate from 47,XXX and 47,XXY. Linden et al. (1995) review the phenotypes of 48,XXXX, 48,XXXY, 48,XXYY, 48,XYYY, 49,XXXXX, 49,XXXXY, 49,XXXYY, 49,XXYYY, and 49,XYYYY. In each, variable intellectual compromise is characteristic, the more so in karyotypes with four or more X chromosomes. While the authors' comment is well taken that the current perception of the seriousness of phenotypic abnormality may have been overstated due to ascertainment bias, and indeed they describe normal (but low) IQs in some of the 2n = 48 karyotypes, it remains true that most have substantial handicap due to intellectual deficit and abnormal behavior (Cammarata et al., 1999). The 49,XYYYY karyotype at PND is reported in Frey-Mahn et al (2003).
XX Male
Discordance of 46,XX karyotype and male genital phenotype, as seen on ultrasonography, allows prenatal diagnosis of the XX male (Ginsberg et al., 1999). Normal intelligence and stature are predicted, but there will be testicular deficiency with infertility (Margarit et al., 1998).
X and Y Chromosome Mosaicism
XX/XY
This is usually pseudomosaicism, resulting from the growth of maternal cells in a 46,XY pregnancy (Worton and Stern, 1984). (Obviously, such pseudomosaicism would normally be undetected if the fetus is female.) Level III XX/XY mosaicism, curiously enough, is most likely to indicate a phenotypically normal female fetus in whom the XY source is unknown, particularly when the XX cells predominate; a male “vanished twin” is a theoretical possibility (Worton and Stern, 1984). A girl born following such a prenatal diagnosis (Hunter et al., 1982) was followed through to mid-adoles-cence, and her development has been entirely normal (A. G. W. Hunter, pers. comm., 2002). Ultrasonography with respect to fetal external genital morphology may be helpful.
A true fetal XX/XY karyotype is rare indeed. Yaron et al. (1999) had a case presenting at amniocentesis, with normal male morphology on ultrasound. The XX/XY mosaicism was confirmed on a second amniocentesis, and, in due course, on the normal male newborn infant (including on genital skin). They could show that that the three X chromosomes all derived from the same maternal X, leading them to suppose a simultaneous X and Y nondisjunction at the second postzygotic division from an initially 46,XY zygote. The abnormal division produced 46,XX and 46,YY daughter cells, with the latter being lost. True hermaphroditism has been recorded only once from an XX/XY amniocentesis result (Amor et al., 1999). The postnatal blood karyotype showed 46,XX[25]/46,XY[5]. Chimerism was presumed. Fetal ultrasonography indicated normal male genital development; at birth, the baby had a short penis with a penoscrotal web, and a normal left testis. At surgery at age 6 months, a right-sided ovary and uterus were seen. Amor et al. note the point that intellectual compromise is not to be anticipated.
X/XY
Patients coming to medical attention and found to have 45,X/46,XY mosaicism range in phenotype from females with classical Turner syndrome, through infants with ambiguous genitalia, to normal but infertile males (Telvi et al., 1999). A risk for gonadal tumor applies (Müller and Skakkebæk, 1990; Müller et al., 1999). By contrast, a normal male infant is the outcome in the considerable majority (90%–95%) of X/XY gestations detected at PND—in other words, cases whose ascertainment was unbi-ased—and going through to birth (Hsu, 1994; Huang et al., 2002). A question might still remain concerning the outlook for fertility. Van den Berg et al. (2000) report a case in which nonmosaic 45,X was diagnosed at short-term CVS, with a nonmosaic 46,XY karyotype seen on long-term culture. Subsequent amniocentesis revealed a true 45,X/46,XY mosaicism. Post-termination, fetal testing showed X/XY mosaicism in all tissues sampled (including gonads). Of 14 pathology studies on fetuses post-termination in Chang et al. (1990), two were found to have ovotestes and one had a “precancerous” lesion.
Other Sex Chromosome Mosaicisms
The karyotypes most frequently seen are 45,X/46,XX, 47,XXY/46,XY, 45,X/47,XXX, and 47,XXX/46,XX. It appears that the considerable majority of cases of true sex chromosomal mosaicism of these types are associated with concordant (Y → male, no Y → female) and normal genital development (Hsu and Perlis, 1984; Wheeler et al., 1988). In X/XX, X/XXX, XXX/X/XX and XXX/XX mosaicism IQ is not discernibly affected; verbal IQ may be slightly lowered in XXY/XY (Netley, 1986; Bender et al., 1993).
X/XX. Koeberl et al. (1995) record 12 cases of 45,X/46,XX mosaicism detected at amniocentesis, with the percentage of 45,X cells in ten of these being in the region of 20%–70%. Postnatal studies (blood and/or skin) confirmed the mosaicism in nine (at a lesser percentage in all but one), while in three of the children no 45,X cells were seen. None showed growth retardation postnatally and in none would a clinical suspicion of Turner syndrome have arisen. Two cases of presumed early ovarian dysfunction, one of these also having urogenital anomalies, might reflect an effect of the karyotype; it is possible some of the remaining cases could also manifest abnormal ovarian function at a later age. The abnormal neurology in one of the twelve is of uncertain significance. As Koeberl et al. commented, “the prenatal diagnosis of 45,X/46,XX is not necessarily benign.” Hsu (1996) and Sybert et al. (1996) debate the validity of previous larger series of X/XX prenatal diagnoses. A clear point forthcoming is that data from longer-term follow-up are desirable, given that a functional gonadal component of the syndrome might not manifest until well into adolescence or adult life.
In 2002, Huang et al. reported their experience with 17 cases of X/XX mosaicism at amniocentesis. The ratios of X to XX cells ranged from 2:23 to 12:3. One case with intrauterine growth retardation (ratio 6:12) terminated in stillbirth, while the remaining 16 had normal ultrasonography. Of the eight cases continuing to term and for which information was available, two liveborn babies had the features of Turner syndrome (ratios 7:10 and 3:14), with the mosaicism confirmed postnatally in one of these. The remaining six (ratios ranging from 3:15 to 12:8) “reportedly had a normal femalephenotype.” To quote Huang et al., “the percentage of 45,X cells in amniocytes does not seem to be an indicator of pregnancy outcome as there was considerable overlap between cases with normal and abnormal outcome.” In a unique case of a monozygous twin pregnancy, one fetus showed nuchal swelling and the other appeared normal (Gilbert et al., 2002). Fetal blood sampling showed low-grade 45,X[2]/46,XX[23] mosaicism in the former, and a normal 46,XX karyotype in the latter, in contrast to postnatal skin fibroblast karyotyping results of nonmosaic 45,X and 45,X[2]/46,XX[78], respectively.
X/XXX/XX and X/XXX. One reported case of X/XXX/XX mosaicism illustrates the difficulty in extrapolating the distribution of cell types from one tissue to another (Schwartz and Raffel, 1992). Amniocentesis gave the proportions 16:20:64, respectively. Cord blood gave similar findings, although in placental tissue (chorion) the percentages were 2:41:57. The baby appeared normal. Huang et al. (2002) reported a case each of X/XXX and X/XXX/XX mosaicism diagnosed at amniocentesis, the former pregnancy producing a newborn with features of Turner syndrome, and the other a normal female. Sybert (2002) reviewed hers and others' data, and concluded that about 60% of girls with X/XXX/XX and X/XXX could be predicted to have short stature, and that “it is fair to suggest that residual ovarian function is possible and to caution that premature ovarian failure is common.”
X/XYY and X/XYY/XY. The X/XYY and X/XYY/XY mosaic states are (necessarily) abnormal in postnatally ascertained cases, but prenatally diagnosed cases have consistently manifested an apparently normal male genital phenotype (Hsu, 1994).
Structurally Abnormal Sex Chromosome
X-Y Translocation. The most common form of the t(X;Y) has the X breakpoint at or distal to Xp22, and the Y breakpoint at Yq11.2 (see p. 117). The intact sex chromosome may be an X or a Y chromosome. With the former, the karyotype is written 46,X,der(X)t(X;Y). A de novo X-Y translocation would be expected to herald a female child, who will likely be short, 150 cm or less in height as an adult (Joseph et al., 1996; Speevak et al., 2001). The site of the breakpoint can be pinpointed with probes for two loci (steroid sulfatase, Kallmann syndrome) in Xp22.3; if these loci are present on the der(X)t(X;Y), intelligence and fertility may be intact, and other defects are unlikely. A few de novo cases have been associated with major defects (Speevak et al., 2001), presumably due to a marginally more proximal Xp breakpoint, with the deletion of crucial genes.
If the intact sex chromosome is the Y, with the karyotype 46,Y,der(X)t(X;Y), the child is expected to be male. If the probes noted above are present, the phenotype is likely to be confined to short stature and infertility. A more extensive loss of loci might determine a nullisomy that would cause important malformation. (X-autosome and Y-autosome translocations are dealt with in the section above on autosomes.)
Other rare types include dicentric X;Y translocations, and der(X) and der(Y) chromosomes with a range of p and q arm breakpoints on X and Y (Hsu, 1994). The phenotypes are male if SRY is present, and otherwise female. Infertility is typical, and, in either sex, short stature. In the der(Y) case, in which there may be an effect of functional X disomy, genital anomaly and other malformation is common, as is mental defect.
X Chromosome Deletion. The possibility of an inherited X;autosome translocation should be checked by doing the mother's karyotype. Alternatively, it may transpire that the mother has the same karyotype. X chromosome deletions in the female, 46,X,del(Xp) or 46,X,del(Xq), predict the probability, but not the certainty, of an incomplete form of Turner syndrome, and/or premature ovarian failure (Fitzgerald et al., 1984; Veneman et al., 1991). Brown et al. (2001)describe a mother, of tall stature (5 feet 10 inches), having a PND of del(X)(q22q26); she had the same karyotype, and “the parents took comfort in the observation that in the mother the deletion had no apparent phenotypic effect.” A normal baby girl was born. Mother and daughter showed completely skewed X-inactivation, the abnormal X being consistently inactive.
In the male, the 46,Y,del(X) state would be nonviable for all but the very smallest deletions, and major abnormality would be probable for those pregnancies that might be viable.
X Chromosome Duplication. De novo X chromosome duplications, 46,X,dup(X), in the female might have been thought to be of minimal effect (a partial XXX syndrome) due to selective inactivation of the abnormal X. This is sometimes but not necessarily the case, and abnormal phenotypes, often including genital defect, are not infrequently observed. Zhang et al. (1996) provide detail according to the extent and site of the duplication in a review of postnatally diagnosed cases. Tihy et al. (1999) describe an infant girl with a de novo dup(X)(q22.1q25) who had physical and neurodevelopmental defects, in spite of the X-inactivation pattern (at least on peripheral blood) showing consistent inactivation of the dup(X). Functional disomy, at least for part of the segment, may contribute to the abnormal phenotype in such cases (Armstrong et al., 2003). Normality, at least at pediatric assessment at 1 month of age, was reported by Lebbar et al. (2002) with respect to an isodicentric X, idic(X)(q27), comprising practically a double copy of the X, identified prenatally, the abnormal chromosome being late-replicating. In the male, functional disomy for the duplicated segment would likely cause severe defects, quite probably lethal in utero.
Other Abnormal X Chromosomes
X chromosome abnormalities are characteristically seen in the mosaic state, the other cell line typically being 45,X. Mosaicism with a large ring X or an isochromosome for the X long arm, 45,X/46,r(X) and 45,X/46,X,i(Xq), respectively, would lead to variant Turner syndrome. An isochromosome for the X short arm, i(Xp), would probably always be lethal, since there would be a functional Xp trisomy (Lebo et al., 1999). As with the tiny ring X syndrome (see following), a marker X that lacks XIST is associated with phenotypic abnormality (Tümer et al., 1998).
The “tiny ring X syndrome” with the karyotype 45,X/46,X,r(X) may have a functional X disomy and is typically, but not universally, seen with a severe phenotype of physical and mental defect, in some resembling Kabuki syndrome. The severity of the phenotype has been attributed to a functional X disomy, due to the ring lacking the XIST locus and thus not undergoing inactivation, although in rare instances this scenario does not apply (Migeon et al., 2000; Stankiewicz et al., 2001d; Tomkins et al., 2002). The phenotype is also expected to be abnormal when the tiny ring is a supernumerary chromosome; in this state it can, very rarely, be seen in the male (Huang et al., 1999a; Le Caignec et al., 2003b).
Y Chromosome Rearrangement
Hsu (1994) lists several possibilities: Yq–deletions of various extents, Yp–, r(Y), i(Yp), idic(Yp), i(Yq), and idic(Yq). Concomitant 45,X mosaicism is often observed, and this can complicate prediction. Absence of SRY leads to female development, and loss of other Yp loci determines a Turner syndrome (TS) phenotype. In 45,X/46,X,der(Y) with TS there is a risk for gonadoblastoma, and since the gonads are inaccessible to easy surveillance, it is generally recommended that they be removed prophylactically (Atkins et al., 2000). Intactness of Yp with loss of Yq loci, and in particular the AZF loci (Figure 5-1), is associated with male infertility; if there is 45,X mosaicism, genital development may be female or ambiguous. The outcomes in one prenatal diagnostic series are set out in Table 25-4.
In the particular case of mosaicism for a 45,X and a 46,X,dic(Y) (isodicentric Y) cell line, the genital sex can be female, male, or ambiguous, and gonadal dysgenesis is common; short stature is frequent (Tuck-Muller et al., 1995; Teraoka et al., 1998). Ultrasonographic assessment of genital anatomy may be helpful. Y locus analysis, including SRY, can be useful, as Hoshi et al. (1998) show in the study of an amniocentesis result of 45,X/46,X,+mar. On FISH analysis, the marker was of Y origin, and most probably an inversion/deletion inv(Y) (q10q11.23),del(Y)(q12). A panel of markers for Yp intervals 1–7 showed intactness for this important functional segment, and the SRY gene was present. In this instance, the child was a normal male, and at age 5 years had normal testosterone levels, making the point that genital defect is not inevitable (inasmuch as can be judged in childhood), as Schwartz et al. (1997) also demonstrate (Table 25-4). Two similar cases with an idic(Y) are described in Huang et al. (2002). This marker may be mitotically unstable in amniocytes, and thus seen, if at all, at only low levels of mosaicism.
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Table 25.4. Outcomes in 11 Cases of Detection at Amniocentesis of an Abnormal Chromosome of X or Y Origin. Note That No Judgment Could Presently be Made Concerning Fertility in the Apparently Normal Males |
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THE “CARRIER FETUS” WHO WILL BECOME A CARRIER ADULT
We have discussed in the introductory chapter the issue of the genetic testing of children. In the case of prenatal diagnosis in which a balanced state is discovered, of course the child has already been tested, and “untesting” is nonsense. Consider the example of the mosaic test result mentioned above, the whole arm translocation 46,XY,t(1;5)(p10;q10)/46,XY. Naturally, parents will want to know what implications this may have for their as yet unborn child. In this example, the genetic risk for the child will be, as the reader can readily determine, essentially that of a likely propensity to miscarriage, should the translocation cell line involve the gonad. It is the counselor's responsibility to communicate this sort of information in outline form to the parents, along with the advice that the child could in the fullness of time attend the clinic on their own behalf. The information must be clearly conveyed. It could be seen as a failure of the counselor's duty of care if, in the next generation, an affected child were born, the parents being unaware of the genetic risk (Burn et al., 1983).
Notes
1. Excepting the unlikely possibility of isozygosity for a recessive gene.
2. An exception may be mosaicism for an isochromosome, as a handful of reports have demonstrated true mosaicism in the context of a single abnormal cell at prenatal diagnosis (see pp. 422).
3. Maternal contamination of the sample may have been the cause (Saura et al., 1998).
4. As noted above, a few instances of apparent nonmosaic trisomy at CVS are also included here, on the assumption that, in the circumstance of a semblance of normal fetal development, a true fetal nonmosaic trisomy for that chromosome would in fact be improbable. We assume in these cases, rather, that this would be either “fetal mosaicism, nonmosaic placenta,” or “fetal–placental mosaicism” with the sampling needle missing the karyotypically normal tissue, each of these scenarios being demonstrated in Figure 25-4. In the specific case of trisomy 16, Yong et al. (2003) note that the outcomes are similar whether the trisomy on CVS is nonmosaic or mosaic.
5. The formally correct nomenclature is actually der(13;13)(q10;q10).
6. This stain specifically distinguishes chromosome 15 from the other acrocentric chromosomes. It also stains qh regions of 1, 9, 16, and Y.
