, . 17:4: 375–379 (1997)

SHORT COMMUNICATION

MISINTERPRETATION OF TRISOMY 18 AS APSEUDOMOSAICISM AT THIRD-TRIMESTER

AMNIOCENTESIS OF A CHILD WITH A MOSAIC46,XY/47,XY,+3/48,XXY,+18 KARYOTYPE

. . . -*, . . . . *, . . . †, . . ‡, . * . . . . *

Departments of *Human Genetics, †Paediatrics, and ‡Obstetrics and Gynaecology, University Hospital Nijmegen,The Netherlands

Received 19 June 1996Revised 30 September 1996Accepted 22 October 1996

SUMMARY

False-negative trisomy 18 has been reported after chorionic villus sampling, but not after amniocentesis. Wedescribe a double aneuploidy in cultured amniocytes that was initially misinterpreted as a pseudomosaicism. Apatient was referred at 31 weeks of gestation because of fetal anomalies at ultrasound examination. Karyotyping ofamniocytes showed a 47,XY,+3 karyotype in 61 clones and a 48,XXY,+18 karyotype in one clone. The latter wasinterpreted as a pseudomosaicism, the more since a second amniocentesis revealed only cells with a 47,XY,+3karyotype. At 36 weeks gestational age, a boy was born with congenital anomalies suggestive of trisomy 18. A bloodculture showed a 48,XXY,+18 karyotype, while in fibroblasts a 47,XY,+3/48,XXY,+18 mosaicism was found.Umbilical cord and bladder epithelial tissue also revealed normal 46,XY cells, besides the aneuploid cells. Therefore,the child proper had a 46,XY/47,XY,+3/48,XXY,+18 mosaicism with the clinical symptoms of trisomy 18. To thebest of our knowledge, this is the first report of a false-negative result of trisomy 18 together with three sexchromosomes after amniocentesis. ? 1997 by John Wiley & Sons, Ltd.

: chromosome mosaicism; trisomy 3; trisomy 18, Klinefelter; double aneuploidy; amniocentesis

INTRODUCTION

Disparity between placental and fetal chromo-some constitutions is a well-known phenomenonthat occurs in 1–2 per cent of all pregnancies(Simoni and Sirchia, 1994). Where chromosomalmosaicism is present in chorionic villus cells butcannot be confirmed in the fetus, this is known asconfined placental mosaicism.A difference between the karyotypes of amnio-

cytes and fetal cells is rare and in most of such cases

a mosaicism is observed in amniocytes, whichincludes the fetal karyotype. In amniocytes, twotypes of mosaicism can be distinguished: truemosaicism and pseudomosaicism (Hsu, 1992).True chromosome mosaicism is defined as an iden-tical aneuploidy in cells from one or more coloniesfrom a minimum of two different culture vessels.Pseudomosaicism can by divided into three differ-ent types: (1) one cell or one region in a colonywith an abnormal karyotype (single-cell pseudo-mosaicism); (2) all the cells of a single colony withan identical aberrant karyotype (single-colonypseudomosaicism); and (3) multiple clones withinthe same culture vessel showing an identical abnor-mal karyotype (multiple-cell pseudomosaicism).

Correspondence to: C. M. A. van Ravenswaaij-Arts, MD,Department of Human Genetics 417, University HospitalNijmegen, P.O. Box 9101, 6500 HB Nijmegen, TheNetherlands.

CCC 0197–3851/97/040375–05 $17.50? 1997 by John Wiley & Sons, Ltd.

In the course of a prenatal chromosome studybecause of intrauterine growth retardation, werecently discovered in cultured amniocytes a singleclone with a double aneuploidy (48,XXY,+18)that was erroneously misinterpreted as pseudo-mosaicism. In all the other amniocyte clonesstudied, trisomy 3 was found. This case reportshows that in some cases it might be presump-tive to consider a chromosome aberration as‘pseudo’-mosaicism, especially when more thantwo different cell lines are involved.

CASE REPORT

A healthy 26-year-old woman was referred to usin the 31st menstrual week of her third pregnancybecause of intrauterine growth retardation. Thefirst two pregnancies had ended in spontaneousabortions at 10 and 7 menstrual weeks, respect-ively. No chromosome studies were performed onthese products, nor on the parents. No ovulation-inducing agents were used in the present pregnancyand no ultrasound evidence of multiple gestationswas observed at the time of referral.Ultrasound investigation revealed additional

polyhydramnios and a bilateral cleft lip in thefetus. Fetal echocardiography was not success-ful. Because a chromosomal abnormality wasconsidered, an amniocentesis was performed.The G-banded preparation of cultured amniotic

fluid cells revealed a 47,XY,+3 karyotype in 61clones obtained from four culture vessels. In oneclone, a 48,XXY,+18 karyotype was seen. Thissingle clone was interpreted as pseudomosaicism.Since a full trisomy 3 has never been described in aliveborn infant, a disparity between the amniocyteand fetal chromosome constitutions was suspected.For further analysis, a cordocentesis was offeredbut refused by the parents.For drainage of the polyhydramnios, a second

amniocentesis was performed at 32 weeks and2·0 l of amniotic fluid was obtained. Karyotypingof this sample again revealed the 47,XY,+3karyotype in eight clones studied.At 38 weeks of gestation labour started and

at the parents’ request, a Caesarean section wasperformed because of fetal distress. A boy wasborn with Apgar scores of 2, 3, and 6 at 1, 5, and10 min, respectively. Birth weight (1746 g), length(41·5 cm), and head circumference (31 cm) were allbelow the 2·3rd centile. Physical examination atbirth revealed hypertelorism, blepharophimosis,

low-set malformed ears, bilateral cleft lip and cleftpalate, micrognathia, small clock-shaped thorax,micropenis, cryptorchidism, and camptodactylyof the fingers and toes (Fig. 1). An ultrasoundinvestigation of the heart showed an atrioventricu-lar septal defect. Echoencephalography revealeddilated ventricles. The phenotype of the childwas very suggestive of trisomy 18. The infantbreathed spontaneously, but needed tube feeding.His clinical condition deteriorated after a few daysand the boy died 10 days after birth, due tocardiorespiratory failure.Post-mortem examination confirmed the cardiac

malformation. In addition, complete aplasia ofboth olfactory nerves and hypoplasia of the caudalpart of the vermis of the cerebellum were found.Cells obtained from the placenta, umbilical cord,

blood, skin, and bladder were karyotyped. Amni-otic cells obtained at the second amniocentesiswere also re-evaluated (Table I).All 129 peripheral blood lymphocytes analysed

had a 48,XXY,+18 karyotype. The skin fibroblastculture showed a 47,XY,+3/48,XXY,+18 mosaic-ism (13/19). Amniotic cells again showed a singleclone with a 48,XXY,+18 karyotype and 44 cloneswith a 47,XY,+3 karyotype.Cells with a normal male karyotype (47,XY)

amongst aneuploid cells were obtained only fromthe bladder and umbilical cord fibroblasts.

DISCUSSION

The present case report illustrates how deceptivethe presence of ‘pseudo’-mosaicism in a prenatalculture can be. The interpretation of mosaic chro-mosome patterns may sometimes be very difficult,not only in chorionic villus cells but also in amnio-cytes. In retrospect, the 48,XXY,+18 clone ofthe first amniocentesis of our patient has beenmisinterpreted as pseudomosaicism, since thechild proper had 46,XY/47,XY,+3/48,XXY,+18mosaicism with the clinical symptoms of trisomy18.The frequency of true chromosome mosaicism

in cultured amniocytes ranges from 0·1 to 0·3per cent (Bui et al., 1984; Hsu and Perlis, 1984;Worton and Stern, 1984). Pseudomosaicisms arefound more often: multiple-cell pseudomosaicismhas a frequency of approximately 1 per cent, whilesingle-cell or single-colony pseudomosaicism isseen in 2–7 per cent of studies. A true chromosomemosaicism with a trisomy 18 or a 47,XXY cell line

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has been diagnosed in amniocytes in at least 17and 59 cases, respectively (Hsu, 1992).In our patient, chromosome studies of the post-

natally obtained biopsies of the chorionic villi weremore in accordance with the karyotype of theinfant than the amniotic fluid culture (Table I).This is an extremely rare situation. False-positiveand false-negative results are obtained far moreoften at chorionic villus sampling than at amnio-centesis. As far as we know, misleading results at

amniocentesis, in contrast to CVS studies, hasbeen described once before. Mascerello et al.(1994) found a mosaic triple trisomy for chromo-somes 13, 20, and 21 at amniocentesis in a karyo-typically normal fetus. Also different biopsies ofthe chorion revealed normal karyotypes.Double autosomal/gonosomal aneuploidy, as

was found in the present case, has been describedbefore, i.e., involving trisomy 8, 18, or 21 (DeBrasiet al., 1995; Jaruratanasirikul and Jinorose, 1994;

Fig. 1—The present patient in frontal (left) and lateral (right) views

Table I—Results of cytogenetic studies in different tissues of the present patient

46,XY 47,XY,+3 48,XXY,+18

Amniotic fluid, 31 weeks (clones) 61 1Blood (cells) 129Skin (cells) 13 19Bladder (cells) 1 5 26Umbilical cord (cells) 12 3 81Chorionic villi, postnatal (cells) 4 60Second examination amniotic fluid,32 weeks (clones) 44 1

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Stevens et al., 1995). Even three cases of a sexchromosome tetrasomy with an additional auto-somal trisomy (8 or 18) have been reported(Sutherland et al., 1978; Kardon et al., 1980; Webbet al., 1984). In double aneuploidy, the super-numerary chromosomes may both be present inthe same cell line, as in our patient. This might becaused by a double non-disjunction event in asingle parent or fertilization of an aneuploidoocyte by an aneuploid sperm. If another (normal)cell line is also present, the aberrant cell line mighthave arisen from a post-zygotic double non-disjunction event. In other cases, two abnormalcell lines, one of which is directly derived from

the other post-zygotically, may be present in thesame individual (i.e., one line with a trisomy 18and the other line with 48,XXY,+18). Finally, twoapparently unrelated aneuploid cell lines may exist(in our patient, a trisomy 3 cell line and a48,XXY,+18 cell line). This situation is rare andseveral alternative mechanisms can be proposedfor the causative events. The most likely expla-nation might be a series of post-zygotic errors(Fig. 2).A full trisomy 3 is found in about 1 per cent of

all early abortuses, whereas a trisomy 3 mosaicismis one of the most common types of mosaicismfound at chorionic villus sampling (Sachs et al.,1990; Guerneri et al., 1989). Most often this rep-resents confined placental mosaicism (Guerneriet al., 1989). At present, we know of only one caseof mosaic trisomy 3 in amniocytes, while a fulltrisomy 3 has not been reported at all (Hsu, 1992).Up to now, four liveborn children, all females,

with a trisomy 3 mosaicism have been described(Metaxotou et al., 1981; Kuhn et al., 1987; Smithet al., 1988; DeKeyser et al., 1988). The number oftrisomy 3 cells in their peripheral leucocytes rangedfrom 3 to 86 per cent. Subsequent fibroblast cul-tures revealed a normal karyotype in two patients.The main features that were shared by all fourpatients were growth retardation and eye abnor-malities. Three patients were mentally retardedand had microcephaly. In Table II the clinicalfeatures of our patient are summarized and

Fig. 2—Diagram showing the possible order of non-disjunctional events, resulting in three different karyotypes

Table II—Clinical features of the present patient compared with patients with a trisomy 3and trisomy 18, respectively

Presentpatient

Trisomy 3n=4

Trisomy 18(%)

Intrauterine growth retardation + 1 96Microcephaly + 2 70Narrow palpebral fissures + 0 80Low-set malformed ears + 2 88Cleft lip palate + 0 20Micrognathia + 1 96Short sternum + 1 68Micropenis/cryptorchidism + n.a. 100Camptodactyly + 1 89Congenital heart defect + 1 85Central nervous system malformation + 0 80

n.a.=not applicable.Data from Metaxotou et al. (1981); Kuhn et al. (1987); Smith et al. (1988); DeKeyser et al. (1988);

Gorlin et al. (1990).

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compared with the features found in the fourpatients with trisomy 3 mosaicism, and in trisomy18 syndrome.To our knowledge, this is the first report of a

false-negative result of trisomy 18 together with agonosomal aneuploidy after amniocentesis. Fur-thermore, we conclude that a full trisomy 3 afteramniocentesis is not to be trusted and needsfurther cytogenetic evaluation.

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