SHORT COMMUNICATION

Prenatal diagnosis and genetic analysis of double trisomy48,XXX,+18

Chih-Ping Chen1,2*, Schu-Rern Chern2, Li-Fan Yeh1, Wen-Lin Chen1, Li-Feng Chen1 and Wayseen Wang2

1Department of Obstetrics and Gynecology, Mackay Memorial Hospital, Taipei, Taiwan, R.O.C.2Department of Medical Research, Mackay Memorial Hospital and National Yang-Ming University, Taipei, Taiwan, R.O.C.

Prenatal diagnosis of simultaneous occurrence of double trisomy involving chromosomes 18 and X isextremely rare. We report on the prenatal diagnosis, genetic analysis and clinical manifestations of a fetuswith both trisomy 18 and trisomy X. A 26-year-old, para 1 woman was referred for genetic counselling at 36weeks' gestation with the sonographic ®ndings of intrauterine growth retardation (IUGR), polyhydramnios,ventricular septal defect, and an enlarged cisterna magna. Both cordocentesis and amniocentesis revealed aconsistent karyotype of 48,XXX,+18. Quantitative ¯uorescent polymerase chain reaction usingpolymorphic small tandem repeat markers speci®c for chromosomes 18 and X rapidly determined thatboth aneuploidies arose as a result of non-disjunction in maternal meiosis II. Our case shows that two non-disjunction events can occur not only in the same parent, but also in the same cell division. Our case alsoshows that double trisomy, 48,XXX,+18, can demonstrate an enlarged cisterna magna, IUGR andpolyhydramnios in prenatal ultrasound. Copyright # 2000 John Wiley & Sons, Ltd.

KEY WORDS: double trisomy; enlarged cisterna magna; meiosis II non-disjunction; trisomy 18; trisomy X

INTRODUCTION

The simultaneous occurrence of double trisomyinvolving chromosomes 18 and X is uncommon. Themajority of affected cases have been reported ininfancy (Uchida and Bowman, 1961; Uchida et al.,1962; Ricci and Borgatti, 1963; Haas and Lewis, 1966;Emberger et al., 1971; Madahar et al., 1974; Vermaand Dosik, 1980; Rosenfeld et al., 1981; Sonoda et al.,1987; Imai et al., 1987; Jaruratanasirikul and Jinorose,1994; Moore et al., 1994; Tsukahara et al., 1994). Toour knowledge, there are very few reports (denHollander et al., 1994; Hanna et al., 1996) concerningprenatal diagnosis of 48,XXX,+18, but the geneticmechanisms by which double trisomy arose were notwell investigated. Here, we present prenatally detecteddouble trisomy for chromosomes 18 and X in a fetuswith an enlarged cisterna magna along with determi-nation of the segregation error responsible for thedouble aneuploidies.

CASE REPORT

A 26-year-old gravida 2, para 1, woman was referredfor genetic counselling in the third trimester because ofabnormal sonographic ®ndings. The mother had onethree-year-old healthy child. At 16 weeks' gestation,she had a maternal serum screening test which showed

a Down syndrome risk of 1 : 5000 calculated from amaternal serum alpha-fetoprotein level of 0.63 multi-ples of the median (MoM) and a total humanchorionic gonadotrophin level of 0.51 MoM. Thepregnancy was uncomplicated until 34 weeks' gesta-tion when intrauterine growth retardation and poly-hydramnios were noted. A level II sonographicexamination at 36 weeks' gestation showed a growth-retarded fetus with a biparietal diameter of 8.6 cm (34weeks), a femur length of 6.2 cm (32 weeks) and anabdominal circumference of 22.1 cm (27 weeks), anamniotic ¯uid index of 38.7 cm, ventricular septaldefect and an enlarged cisterna magna measuring19 mm in anteroposterior dimension. Genetic amnio-centesis and cordocentesis were immediately per-formed. Both amniocytes and blood lymphocyteshad a consistent karyotype of 48,XXX,+18(Figure 1). Quantitative ¯uorescent polymerase chainreaction (QF-PCR) using polymorphic small tandemrepeat (STR) markers speci®c for chromosomes 18 andX rapidly determined double trisomy for chromo-somes X and 18 and the maternal origins of theaneuploidies (Table 1, Figure 2). A female baby(Figure 3) was subsequently delivered with clinicalmanifestations of low-set ears, clenched hands, rocker-bottom feet, micrognathia, prominent occiput, hypo-tonia, ventricular septal defect, patent ductusarteriosus, cerebellar hypoplasia and an enlargedcisterna magna, but without evidence of renal abnor-mality. Birth weight was 1230 g (<3rd percentile) andlength was 36 cm (<3rd percentile). Apgar scores were4 and 8 at 1 min and 5 min, respectively. The infantdied of cardiopulmonary failure two weeks after birth.

*Correspondence to: C.-P. Chen, Department of Obstetrics andGynecology, Mackay Memorial Hospital, 92, Section 2, Chung-Shan North Road, Taipei, Taiwan, Republic of China.E-mail: cpc-mmh@yahoo.com

PRENATAL DIAGNOSIS

Prenat Diagn 2000; 20: 750±753.

Copyright # 2000 John Wiley & Sons, Ltd. Received: 4 February 2000Revised: 22 May 2000

Accepted: 28 May 2000

Microsatellite analysis

Fetal DNA was isolated from cord blood and parentalDNA from peripheral blood using a DNA extractionkit (Qiagen, Hilden, Germany). QF-PCR and STRmarkers were used for rapid determination of aneu-ploidy and parental origin of the extra chromosomes.Seventeen pairs of primers for the polymorphicmarkers, i.e., D18S59, D18S378, D18S51, D18S386,D18S70, D18S542, D18S843, D18S847, D18S974,D18S977, DXS987, DXS981, DXS101, DXS1001,DXS1189, DXS1356, and DXS6789 were used forPCR ampli®cation (Internet Web of STRs, 1999).

The PCR reaction was carried out in a 10 ml reactionmixture containing 30 ng DNA, 0.33 mM of eachprimer, 0.25 mM dNTPs, 1rPCR buffer, 1.5 mM

MgCl2, and 0.5 U AmpliTaq (Perkin Elmer, Norwalk,USA). The following thermal pro®le was followed.After initial denaturation at 95uC for 5 min, 32 cyclesof PCR ampli®cation were done: 45 s denaturation at94uC, 45 s annealing at 55uC, and 55 s extension at72uC. The ®nal extension was at 72uC for 15 min. TheDNA fragments were diluted 15 times (HEX-labelledproducts) or 20 times (6-FAM- and TET-labelledproducts) and were mixed together with formamideand Genescan-500 TAMRA size standard (AppliedBiosystems, Foster City, USA). The DNA fragmentswere resolved on a 4% denaturing polyacrylamide geland analysed by an automated DNA sequencer (ABIPrism 377, Applied Biosystems, Foster City, USA) anda Genescan Analysis 2.1 software (Applied Biosys-tems, Foster City, USA). The amounts of amplicons

Figure 1ÐKaryotype of the proband

Table 1ÐGenotypic information of the fetus and parents at STR markers speci®c for chromosomes 18 and X obtained byQF-PCR assaysa

Marker Chromosome location Father Mother Fetus

D18S378 18pter-p11.2 264, 268 260, 264 264, 264, 268D18S386 18q22.1-q22.2 340, 348 348, 352 340, 352, 352D18S51 18q21.3 275, 283 283, 287 283, 287, 287D18S542 18 182, 186 182, 186 182, 182, 186D18S59 18pter-p11.22 149, 155 157, 159 149, 159, 159D18S70 18q23 115, 119 113, 121 115, 121, 121D18S843 18 185, 191 182, 182 182, 182, 185D18S847 18 220, 232 216, 224 216, 216, 220D18S974 18 258, 258 262, 262 258, 262, 262D18S977 18 253, 261 253, 265 253, 265, 265DXS101 Xq22 191 194, 194 191, 194, 194DXS1001 Xq24 210 208, 214 210, 214, 214DXS1189 X 231 223, 231 223, 223, 231DXS1356 X 219 215, 223 219, 223, 223DXS6789 X 160 168, 168 160, 168, 168DXS981 Xq11.2-q13.1 190 190, 190 190, 190, 190DXS987 Xp22 214 214, 218 214, 218, 218

aAlleles (basepair sizes) are listed below each individual.

PRENATAL DIAGNOSIS OF DOUBLE TRISOMY 48,XXX,+18 751

Copyright # 2000 John Wiley & Sons, Ltd. Prenat Diagn 2000; 20: 750±753.

were estimated based on the peaks on the electro-phoretograms. The ampli®cation products were sizedaccurately.

Table 1 shows allelic distribution of STR markersspeci®c for chromosomes 18 and X in the fetus andparents. The specimen showed a diallelic pattern witha dosage ratio of 2 : 1 for chromosome 18- and X-speci®c markers. The fetus inherited two copies of asingle maternal allele in chromosome 18 and chromo-some X. The molecular result, along with thekaryotype of 48,XXX,+18, was consistent with theoccurrence of double non-disjunction in maternalmeiosis II.

DISCUSSION

Molecular genetic analyses using polymorphic DNAmarkers have enabled the study of non-disjunction offetal aneuploidy. Recently, QF-PCR and STR markershave been used for rapid detection of fetal aneuploidyas well as determination of parental origin and geneticoccurrence at meiosis I or II (Adinol® et al., 1997).This method was useful in our investigation ofparental origin and the stage of meiotic error in thepresent case of which segregation studies of twochromosomes were required.

Most cases of double aneuploidies in livebornsinvolve the sex chromosomes combined with eithertrisomy 13, 18, or 21; i.e., XXX/18, XXX/21, XXY/13,XXY/18, XXY/21, XYY/13, XYY/18 and XYY/21(Borgaonkar, 1997; Reddy, 1997). Other rare combi-nations include 13/18, 13/21, 18/21, 21/21, and XXYY(Borgaonkar, 1997; Reddy, 1997). In spontaneousabortions, the double aneuploidies often involve at

least one non-viable trisomy such as 16, 8, 15, or 2,and the frequent combinations include 16/21, 2/16,15/16, 2/18, 8/16, 13/16 and 16/22 (Reddy, 1997). Ourcase was born alive and involved two viable aneuploi-dies XXX/18.

Studies of the extra chromosome in trisomy 18 haverevealed that 91% are of maternal origin (Nicolaidisand Petersen, 1998), 60% are due to an error at meiosisII, 31% are due to an error at meiosis I, and about 8%are due to mitotic error (Fisher et al., 1995;Eggermann et al., 1996; Nicolaidis and Petersen,1998). On the other hand, 93.5% of 47,XXX resultfrom maternal non-disjunction at meiosis I (Robinsonand de la Chapelle, 1997). In this presentation, bothtrisomy 18 and trisomy X arose as a result of non-disjunction in maternal meiosis II. Our case shows thattwo non-disjunction events involving both chromo-somes 18 and X can occur not only in the same parent,but also in the same cell division. Park et al. (1995)described a similar segregation error in a fetus with48,XXX,+21 and suggested a general cellular defect,such as impaired spindle function or improper signal-ling of sister chromatid segregation.

Infants with 48,XXX,+18 can manifest typicalabnormalities of trisomy 18 such as intrauterinegrowth retardation (IUGR), typical facial appearance,congenital heart defects, overlapping and ¯exiondeformity of ®ngers, rocker-bottom feet and renalabnormality (Jaruratanasirikul and Jinorose, 1994).Reported abnormal ultrasound ®ndings include poly-hydramnios, nuchal thickening, hydrocephaly, tra-cheo-oesophageal ®stula and omphalocele (Hannaet al., 1996). The potentially detectable trisomy 18fetuses may escape early prenatal screenings anddemonstrate an enlarged cisterna magna in lategestation (Chen et al., 1998). The 47,XXX femaleshave a higher than normal incidence of minorstructural abnormalities such as epicanthic folds, earanomalies and clinodactyly (Robinson and de laChapelle, 1997). Our case shows that fetuses with48,XXX,+18, like trisomy 18, can demonstrate anenlarged cisterna magna, IUGR and polyhydramniosin prenatal ultrasound. The ability of accuratediagnosis of fetal anomalies before birth has expanded

Figure 2ÐRepresentative electrophoretograms for paternal blood(A), maternal blood (B), and cord blood (C)

Figure 3ÐThe proband at birth

C.-P. CHEN ET AL.752

Copyright # 2000 John Wiley & Sons, Ltd. Prenat Diagn 2000; 20: 750±753.

perinatal management options. We emphasize thatprenatal sonographic detection of an enlarged cisternamagna associated with IUGR and polyhydramniosin late gestation should prompt rapid genetic diag-nosis and a detailed search for other structuralabnormalities.

ACKNOWLEDGMENT

This work was supported by a research grant(NSC-89-2314-B-195-011) from the National ScienceCouncil, Taiwan, R.O.C.

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PRENATAL DIAGNOSIS OF DOUBLE TRISOMY 48,XXX,+18 753

Copyright # 2000 John Wiley & Sons, Ltd. Prenat Diagn 2000; 20: 750±753.