European Human Genetics Conference 2007 June 16 – 19, 2007 ...

Prenatal diagnosis
P0438. Is fetal DNA detectable in maternal urine?
S. Majer, M. Bauer, I. Oreskovic, A. Strele, E. Giegerl, M. Eder, U. Lang, B.
Pertl;
Medical University of Graz, Graz, Austria.
Objectives: The aim of this study was the detection, quantification, and
correlation of cell-free fetal (cff) DNA in maternal urine and plasma in
normal and complicated pregnancies during the third trimester.
Methods: 151 urine and plasma samples obtained from 96 women
pregnant with male and 55 pregnant with female fetuses were collected
and analyzed for cff-DNA using fluorescent PCR and quantitative
real-time PCR. The concentrations of cff and total DNA in maternal
plasma were correlated with maternal and obstetric parameters using
appropriate correlation analyses.
Results: Y-chromosome specific sequences were detected in 31/96
(32.3%) urine samples collected from women pregnant with male fetuses
using the DYS14-assay and in 6/96 (6.3%) urine samples using
the SRY-assay for real-time PCR analysis. DNA was extracted from
1 ml maternal urine using the QIAamp®MinElute Virus Spin Kit (QIA-
GEN). No cff-DNA was detected in all 55 urine samples obtained from
women pregnant with female fetuses. All 96 plasma samples obtained
from women pregnant with male fetuses were tested positive for cff-
DNA using real-time PCR. Cff-DNA exhibited a correlation with gestational
age (R=0.244; P=0.018) and an inverse correlation with the
latency between blood collection and birth (R=-0.218; P=0.036). Total
DNA showed a correlation with placental weight (R=0.182; P=0.034)
and pregnancies associated complications (R=0.280; P35-year-old)
and 4 of them had an increased maternal serum screening test result.
Of 95 cases, 86 had isolated CPC. Nine of them had secondary ultrasonographic
abnormality such as intracardiac ecogenic focus, cardiac
hypertrophy, clinodactyly, renal pelvis dilatation, hydronephrosis, hyperecogenic
kidney, ventriculomegaly and overlapping fingers. While
cytogenetic investigation revealed karyotypic abnormality in 8 fetuses
(8.42%), 88 were normal (41 fetuses were 46,XX and 47 were 46,XY).
Among the abnormal karyotypes, 3 fetuses (3.16%) presented Trisomy
18. The other abnormal karyotypes were 46,XY[144]/47,XY+21[6],
46,XY,inv(9)(p11;q13) (2 fetuses) and 46,XY,15p+.
P0440. Fluorescence in situ Hybridization in prenatal diagnosis
H. Ljubic, R. Lasan, L. Letica, M. Burek, D. Muzinic, D. Begovic;
Division of Genetics and Metabolism, Department of Pediatrics, University
Hospital Centre, Zagreb, Croatia.
Samples of amniotic fluid from 49 patients were analyzed using Fluorescence
in situ Hybridization (FISH) technique, applied on cultured
and uncultured amniotic cells. Indications for FISH analysis on uncultured
amniotic cells were: late weeks of gestation, abnormal ultra-
1 2
sound markers, abnormal serum-screening test, chromosome reciprocal
translocation in parents and sex-chromosome related diseases.
Analyses on 17 samples of uncultured amniocytes revealed two pathological
karyotypes (11,8 %). FISH analysis on 32 cultured samples
was used as adjunct technique after classical cytogenetical analysis.
Indications for 24 samples were suspected chromosomal structural de
novo and numerical aberrations, 5 were investigated for population
variants of acrocentric chromosomes and 3 for parents carriers of reciprocal
translocation.
From total number of 32 samples, 14 karyotypes were pathological
(43,8 %) and 18 karyotypes were normal (56,2 %).
Following analyses on cultured amniocytes, 4 medically indicated
abortions were recommended as well as 1 medically indicated abortion
after uncultured amniocytes analysis.
FISH is a fast, powerful molecular cytogenetic technique used as an
supplement to conventional chromosomal analysis.
P0441. Prenatal diagnosis of chromosomal abnormalities in
Lithuania
N. Krasovskaja 1 , E. Benusiene 1,2 , E. Butkeviciene 1 , V. Kucinskas 1,2 ;
1 Center for Medical Genetics, Vilnius, Lithuania, 2 Department of Human and
Medical Genetics, Faculty of Medicine, Vilnius University, Vilnius, Lithuania.
Objective. To ascertain the effectiveness of non-invasive prenatal testing
of chromosomal abnormalities.
Methods. The risk for chromosomal defect was calculated by taking
in account maternal age and gestation, first and/or second trimester
serum biochemistry and first and/or second trimester ultrasound. The
vast majority of women were tested in the second trimester. Cut-off
for invasive testing was risk 1 in 250. For diagnosis karyotype analysis
from cultured amniocytes and quantitative fluorescent polymerase
chain reaction was performed.
Results. 827 amniocenteses were performed during the period of 4,5
years . Abnormal karyotypes were ascertained in 5.56% of cases. The
structure of abnormalities was: trisomy 21-47.8%, trisomy 18-26%,
mosaic karyotypes-8.7%, Robertsonian translocations-6.5%, reciprocal
translocations-2.17%. In the group of women after non-invasive
testing was born 5 babies with trisomy 21. The fetal loss after invasive
procedures was 0.96%.
Conclusions. The effectiveness of non-invasive prenatal diagnostics of
trisomy 21 was 81.5%. The vast majority of non-invasive and invasive
tests should be performed in the first versus the second trimester of
pregnancy.
P0442. Prenatal molecular diagnosis of Cockayne Syndrome
C. Conte, M. D’Apice, F. Sangiuolo, A. Botta, G. Novelli;
Dpt Biopathology, Rome, Italy.
Cockayne Syndrome (OMIM 216400) is a rare and multi-systemic recessive
disease characterized by postnatal growth failure and progressive
multiorgan dysfunction. Growth and developmental abnormalities
become fully manifest in the second year of life. Progressive impairment
of clinical features leads to severe disability, and death occurs
within the first or second decade of life. CSA is caused by mutations in
the 12 exons of ERCC8 gene, encoding a protein involved in the group
of excision-repair cross-complementing process. Upnow 6 mutations
has been characterised in the CSA gene. In our laboratory we examined
a family with an affected son, died at the age of twelve, presenting
typical clinical features. At the age of seven he weighed 9.360 gr and
he was 48 cm high. The proband resulted compound heterozygote
for two new mutations: a deletion del1436-586 (segregating from the
mother) detectable by RNA analysis and a missense mutation C336G
located in exon 4 (from the father). Southern analysis was performed
to characterised the extension of the deletion. Successively the mother
undergoes prenatal diagnosis by transabdominal chorionic villus sampling
(CVS) at week XII of pregnancy. Direct analysis on DNA and RNA
was performed. A linkage analysis was also developed to study the
segregation of the haplotypes, using markers located within an interval
of 2.5cM comprising ERCC8 gene. This study allowed us to identify a
foetus with a wild genotype confirmed at the birth by analysis on biological
material isolated from placenta. This study documents the first
molecular prenatal diagnosis of Cockayne syndrome.