Prenatal diagnosis of trisomy 9 mosaicism: Two new cases

We present two prenatal cases of trisomy 9 mosaicism, both of which presented intrauterine growth retardation (IUGR) and other abnormal ultrasound findings. In case A, mosaicism was found in amniotic fluid cell cultures, of which 65 per cent were trisomic cells, on average. In case B, trisomic cells were present in amniotic fluid cell cultures (12 per cent) but none were found in fetal cord blood. After autopsy, cytogenetic findings were confirmed in different tissue cultures. It is concluded that echographic indicators are a very useful tool for a correct prenatal diagnostic interpretation of trisomy 9. Suspected trisomy 9 mosaicism always requires further investigation and fetal cord blood cytogenetic analysis may not be considered as providing an accurate diagnosis of fetal trisomy 9.     

Prenatal diagnosis of trisomy 9 mosaicism possibly limited to fetal membranes

In repeat amniotic fluid cultures mosaicism due to trisomy 9 was noted. Autopsy of the aborted female fetus showed a sinus urogenitalis and gonadal dysgenesis with absence of germ cells only. Fetal lymphocytes and skin fibroblasts had a normal karyotype but trisomy 9 was found in cells grown from placenta. It is likely that trisomic cells were limited to fetal membranes.     

Trisomy 20 mosaicism in amniotic fluid cells

The significance of trisomy 20 mosaicism in cultured amniotic fluid cells is still confusing. We report a case of amniotic cell normal/trisomy 20 mosaicism diagnosed prenatally. The pregnancy was carried to term and a normal baby girl was delivered. The authors consider that in cases of amniotic fluid cell normal/trisomy 20 mosaicism the termination of pregnancy may not be advised, however, the parents should be fully informed.     

An infant with trisomy 9 mosaicism presenting as a complete trisomy 9 by amniocentesis

We present a case in which amniocentesis performed at 33 weeks' gestation because of symmetrical intrauterine growth retardation and decreased amniotic fluid volume led to the prenatal diagnosis of a fetus with a karyotype of 47,XX,+9, t(1;20)(q42;p11.2) pat, i.e., with an extra chromosome 9 and a balanced translocation between chromosomes 1 and 20. At delivery, the baby showed clinical features of trisomy 9, yet chromosome analysis of the cord blood revealed no trisomy 9 cells, a finding confirmed by neonatal blood karyotyping. The balanced translocation was present in all cells. A skin biopsy confirmed trisomy 9 mosaicism with 10 per cent trisomy 9 cells. The baby died at 6 weeks and an autopsy was obtained. Chromosome analysis of different organs demonstrated different frequencies of the mosaicism of trisomy 9. The possible underlying mechanism for the discrepancy between the karyotype results by amniocentesis and those of other tissues is discussed.     

Trisomy 12 mosaicism detected by mid–trimester amniocentesis

Trisomy 12 mosaicism was found in about 15 per cent of cultured amniocytes obtained from a 32-year-old white female at 17·6 weeks of gestation. Termination of pregnancy was elected and multiple tissues were obtained for chromosome analysis. Of 158 cells examined, only 1 cell in placenta was found with an extra number 12 chromosome. Pathological examination of the fetus did not reveal significant physical abnormalities. This report illustrates the difficulty of confirming trisomy 12 mosaicism which has been detected on prenatal diagnosis. The presence of trisomy 12 in one placental cell obtained from the curettage specimen suggests the possibility of confined placental mosaicism in this case.     

Trisomy 5 mosaicism detected prenatally with an affected liveborn

This paper reports a case of chromosomal mosaicism for trisomy 5 recovered from amniotic fluid cells and from skin fibroblasts of a liveborn dysmorphic male. Routine amniocentesis was performed at 16 weeks' gestation because of parental concern. Trisomy 5 cells were measured from 25 per cent of amniocytes from two culture vessels. No further invasive testing was performed until 32 weeks' gestation, at which time ultrasound examination showed fetus with intrauterine growth retardation. Fetal blood sampling was then performed, with only karyotypically normal cells recovered. At birth, the child was found to have multiple dysmorphic features and congenital anomalies, including an eventration of the diaphragm and ventricular septal defect, both of which required surgical correction. Chromosomal analysis of cord blood lymphocytes indicated 46,XY; however, 20 per cent of the cultured fibroblasts obtained from the chest skin at the incision site for diaphragmatic repair had a 47,XY,+5 karyotype. Trisomy 5 mosaicism may be another example of tissue-limited mosaicism. Fetal blood sampling can then be falsely reassuring. Furthermore, because some cell lines rarely appear in lymphocyte populations, cytogenetic analysis of multiple tissues warranted as part of the evaluation of individuals with developmental delay and dysmorphic features.     

Prenatal detection of trisomy 9 mosaicism

Chromosomal mosaicism in amniotic fluid cells poses a serious dilemma in prenatal diagnosis since the observation may represent: (1) pseudomosaicism—an inconsequential tissue culture artefact; or (2) true mosaicism—occurring in approximately 0.0 per cent of amniocenteses with a significant impact on pregnancy outcome. Mosaicism for trisomy 9 was observed in an amniotic fluid specimen obtained for advanced maternal age with two cell lines [46,XX (46 per cent)/47,XX, + 9 (54 per cent)] present in each of four culture flasks. Since more than 75 per cent of newborns with trisomy 9 mosaicism have complex cardiac malformations, a fetal echocardiogram was obtained at 20 weeks' gestation and interpreted as normal. A fetal blood sample (22 weeks' gestation) disclosed only a single trisomy 9 cell among the 100 metaphases analysed. However, a second fetal echocardiogram performed at the time of blood sampling suggested a non-specific cardiac anomaly. Fetal autopsy following elective pregnancy termination revealed several malformations including severe micrognathia, persistence of the left superior vena cava, and skeletal anomalies. Cytogenetic studies of cell cultures derived from several fetal tissues demonstrated trisomy 9 ranging from 12 to 24 per cent.     

Parental mosaic trisomy 21 detected following maternal cell contamination of an amniotic fluid specimen from a normal male pregnancy

We report a case of maternal mosaic trisomy 21 ascertained at prenatal diagnosis as a result of maternal cell contamination of an amniotic fluid sample. A 34 year old female was referred for karyotyping because of a previous trisomy 21 pregnancy. Chromosome analysis of primary in situ cultures showed a karyotype of 47,XX, + 21[6]/46,XY[32]/46,XX[2]. Molecular testing demonstrated maternal cell contamination of the amniotic fluid sample and G-banded karyotyping of maternal blood showed that 3/200 cells had trisomy 21, consistent with the mother being a Down syndrome mosaic. A normal male baby with a 46,XY chromosome complement was delivered at 30 weeks. This case emphasises the need for close collaboration between cytogenetic and molecular genetics laboratories in resolving unusual cases of mosaicism. Copyright © 2007 John Wiley & Sons, Ltd.     

Trisomy 20 mosaicism

A further case of trisomy 20 mosaicism found at amniocentesis is presented. Pregnancy was terminated, the fetus showed facial dysmorphia and minor cardial and renal anomalies. 19 published reports of true trisomy 20 mosaicism at amniocentesis are reviewed. Five pregnancies resulted in obviously normal newborns. The significance of mostly minor anomalies found at autopsy of 7 fetuses remains unclear. With regard to genetic counselling the significance of trisomy 20 mosaicism is summarized as follows: (1) true trisomy 20 mosaicism in amniotic fluid cells reflects mosaicism of the fetus; (2) severe malformation is not a major feature of trisomy 20 mosaicism; (3) the risk of mental retardation is still undetermined, due to limited experience. However, there is no definite proof that the condition is harmful at all.     

Management of prenatally detected trisomy 8 mosaicism

We report on ten pregnancies with trisomy 8 mosaicism. Nine cases were prenatally detected in chorionic villi (n=6), amniotic fluid (AF) cells (n=2) or fetal blood (FB) lymphocytes (n=1). Follow-up laboratory investigations showed confined placental mosaicism (CPM) or pseudomosaicism in eight cases. In one case with ultrasound abnormalities, trisomy 8 mosaicism was detected in FB cells although cultured AF cells showed normal cells only. Another case of mosaic trisomy 8 was prenatally missed; cytogenetic analysis of short-term cultured villi revealed a normal male karyotype, while postnatally, trisomy 8 mosaicism was detected in peripheral blood lymphocytes and skin fibroblasts of the affected child. These findings indicate the difficulties in the prenatal diagnosis of trisomy 8 mosaicism. When found in chorionic villi, it mostly represented CPM, while in a case of true fetal trisomy 8 mosaicism, the cytotrophoblast cells showed a normal karyotype. So, the cytotrophoblast compartment of chorionic villi is a poor indicator of the presence or absence of fetal trisomy 8 mosaicism. Follow-up investigations including amniocentesis and especially fetal blood sampling are required to come to a definite prenatal diagnosis of trisomy 8 mosaicism. Copyright © 2001 John Wiley & Sons, Ltd.     

Prenatal diagnosis of mosaic trisomy 9

Mosaic trisomy 9 was detected in an amniotic fluid cell culture from a 40-year-old woman evaluated because of advanced maternal age. After counselling, parents elected to terminate the pregnancy. On autopsy the fetus was found to have hydrocephalus and a single kidney. The diagnosis of trisomy 9 mosaicism was confirmed in cultured skin fibroblasts. This is the third reported case of trisomy 9 mosaicism diagnosed prenatally.     

Filtration and recirculation of early amniotic fluid. Evaluation of cell cultures from 100 diagnostic cases

Due to the low cell concentration, cultures from early amniotic fluid specimens usually require 2–3 weeks in culture prior to karyotyping. The purpose of this study was to evaluate the culture quality of amniotic fluid cells from early pregnancy, obtained by a new filter technique. The hypothetical advantage of the technique was that the increased cell yield might reduce the culture time before karyotyping. Culture quality was assessed by the number of colonies, the percentage of colonies containing mitoses in filter and control cultures, and the culture time. The setting was a consecutive clinical trial. One hundred samples were obtained from ongoing pregnancies at 11–14 weeks of gestation (mean 12·8 weeks). By circulating a mean of 26 ml of amniotic fluid through a cell filter system leading the cell-free fluid back to the amniotic cavity, the cell yield was increased in the sample of 7 ml corresponding to the dead space of the filter system. The culture results were compared with control cultures from 5 ml samples drawn from the same pregnancies prior to recirculation. The cultures from the first flushing of the filter system yielded 2·6 times more colonies and in total 4·2 times more colonies were found in the three cultures grown from each filter sample when compared with the control cultures. Moreover, the filter cultures showed significantly more colonies with mitoses. The mean culture time was 8·0 days for the filter cultures, from which the karyotypes were analysed. The controls would have needed more time in culture to fulfil the diagnostic criteria for karyotyping. One case of 47,XY, + 21 was found; the rest had normal karyotypes. We conclude that the filter technique improves the culture quality of early amniotic fluid samples and allows early arrest of the cultures.     

Resorbed co-twin as an explanation for discrepant chorionic villus results: Non-mosaic 47,XX, +16 in villi (direct and culture) with normal (46,XX) amniotic fluid and neonatal blood

Non-mosaic trisomy 16 was observed in chorionic villus cytotrophoblasts (direct) as well as cultured mesenchymal core cells derived from the pregnancy of a 38-year-old woman. Chromosome preparations from amniotic fluid and neonatal cultures (cord blood) were 46,XX. Normal fetal growth as determined by serial ultrasound examinations occurred throughout the pregnancy, which resulted in a healthy 2724 g female. Multiple biopsies taken from the umbilical cord, placental cotyledons, and fetal membranes were 46,XX. However, a placental nodule and three of six cultures initiated from membranes (amnion and chorion) showed 46,XX/47,XX, + 16 mosaicism. We propose that the trisomy 16 cells arose from residual villi derived from a trisomic cotwin that never developed. This case further demonstrates that normal fetal growth may presage normal outcome irrespective of cytogenetic findings in cytotrophoblasts (direct) and cultured mesenchymal core cells.     

Mosaic trisomy 17 in amniotic fluid cells not confirmed in the newborn

A case of true fetal mosaicism 46,XY/47,XY, + 17 was diagnosed in amniotic fluid cells. After genetic counselling and unsuccessful periumbilical blood sampling the pregnancy continued to term, and a healthy male infant was born. Lymphocytes of the newborn had a normal karyotype. Follow-up of the child at age 18 months showed normal physical and mental development indicating that the trisomic cell line was restricted most probably to the extra fetal tissue.     

Prenatal diagnosis,fetal pathology and cytogenetic analysis of A 46,XX/47,XX, + 15 mosaic

Mosaic trisomy 15 was prenatally diagnosed on amniotic fluid cells from two consecutive amniocenteses and was confirmed on cells from five different fetal tissues. The proportion of normal versus trisomic cells was consistently higher in the amniotic cell cultures and-with one exception-in the fetal tissues, while serial subcultures gave different results. The slightly atypical external features and internal malformations of the affected fetus as compared to the only clinical observation from the literature are not unusual enough to allow the delineation of a specific malformation pattern.     

The dilemma of a low rate of chromosomal mosaicism found in fetal blood sampling

We report a case in which mosaicism of trisomy 13 was detected in 4/10 cells (40 per cent) in amniotic fluid cell cultures, followed by a low rate of mosaicism (1/160 cells) detected in a fetal blood sample. This finding presents a dilemma both for the genetic counsellor and for the parents in determining whether or not to terminate the pregnancy.     

Mosaicism for trisomy 12: Four cases with varying outcomes

Trisomy 12 observed in chorionic villus sampling (CVS) may reflect generalized mosaicism or indicate mosaicism confined to only the placenta. In this report, four cases of trisomy 12 observed in CVS or cultured placental biopsies with varying outcomes are presented. Seven dinucleotide repeat polymorphisms for chromosome 12 were used to determine the chromosome 12 origins in the fetus or child and to delineate the mechanism(s) that gave rise to the trisomy. In two cases (cases A and C), the mosaicism was confined to the placenta, resulting in normal liveborns. Although, in one case, the molecular results suggested an apparent duplication of one paternal chromosome 12 in the placenta, normal biparental inheritance was found in the diploid fetal cell line in both cases. In two other cases (cases B and D), trisomy 12 was observed in both extraembryonic and fetal tissues. In one of these pregnancies, a child was born by Caesarean section at 37 weeks because of intrauterine growth retardation and oligohydramnios, and resulted in neonatal death. Molecular markers and fluorescence in situ hybridization (FISH) revealed low-level trisomy 12 mosaicism in the spleen. In the fourth case, fetal abnormalities were detected on ultrasound and low-level trisomy 12 mosaicism was observed in amniotic fluid cells using conventional cytogenetics and FISH. Molecular markers revealed a maternal meiosis I non-disjunction of chromosome 12 in DNA from a cultured placental biopsy. Although predicting the outcomes of pregnancies involving confined placental mosaicism remains difficult, molecular techniques are valuable tools for distinguishing uniparental from biparental disomy and mechanisms of mosaicism.     

Alkaline phosphatase activity in amniotic fluid in pregnancies with fetal disorders

Alkaline phosphatase (ALP) activity was determined in 255 amniotic fluid samples collected by amniocentesis between 15 and 39 weeks of gestation. The samples were originally used for chromosomal analysis and/or alpha-fetoprotein measurements. The mean ALP activity in early amniotic fluid from pregnancies with fetal trisomy 18 and 21 syndromes was half of that found in the controls. Highly elevated ALP activity (over 10 times the median level) was found in 14 samples. Two of these pregnancies had normal outcome. Three samples were from pregnancies with intrauterine fetal death. Fetal disorders, including abdominal wall defect (four cases), Meckel's syndrome (two), hydrops fetalis syndrome (two) and genital anomaly (one), were observed in nine cases. Moderately elevated ALP activity (over three times the median) was found in 10 cases, including five pregnancies with a preterm labour shortly after the sample collection. The results indicate that elevated ALP activity in the third trimester amniotic fluid is often associated with fetal disorders.     

Trisomy 20 mosaicism confirmed in a phenotypically normal liveborn

Prenatal diagnosis of trisomy 20 mosaicism in this case was based on cytogenetic analysis of cultured amniotic fluid cells (23/52 cells were trisomy 20 representing cells from each of four primary cultures). The pregnancy continued to term and the mosaicism was confirmed in the phenotypically normal male neonate by analysis of cultured foreskin fibroblasts (7/49 cells + 20) and placental cells 20/20 cells + 20) whereas the peripheral lymphocytes were cytogenetically normal (20/20 cells were 46,XY). This represents the first confirmation of trisomy 20 mosaicism in a phenotypically normal full-term neonate.     

Genetic amniocentesis in biamniotic twin pregnancies by a single transabdominal insertion of the needle

We present a technique to aspirate amniotic fluid from both sacs in biamniotic twin pregnancies using a single abdominal insertion with a spinal needle. It was successful in 48 out of 55 cases of biamniotic twin pregnancies referred to our perinatal unit between 1985 and 1994. The single insertion technique was used when the inter-amniotic membrane was clearly evident and two separate free amniotic fluid pools could be reached by the operator with a single puncture. An adequate amount of amniotic fluid was sampled from both sacs to make a cytogenetic diagnosis in all cases. There were four fetuses with trisomy 21 in three twin pregnancies. In two cases, only one twin was affected whilst the co-twin was normal, so that a selective feticide was performed. No miscarriages due to genetic amniocentesis were reported. After 1990, all genetic amniocenteses in biamniotic twin pregnancies (except for one case due to late booking) were performed between 14 and 15 weeks of gestation and with all cases except one, it was possible to sample both twins by a single puncture. We suggest that early amniocentesis (14–15 weeks) by a single abdominal puncture could be a reliable and safe alternative to first-trimester chorionic villus sampling in twin pregnancies.