Pallister-Killian syndrome: Cytogenetics and molecular investigations of mosaic tetrasomy 12p in prenatal chorionic villus and in amniocytes. Strategy of prenatal diagnosis.

OBJECTIVE: Pallister-Killian syndrome (PKS) is a rare, sporadic genetic disorder caused by mosaic tetrasomy of the short arm of chromosome 12 (12p). Clinically, PKS is characterized by several systemic abnormalities, such as intellectual impairment, hearing loss, epilepsy, hypotonia, craniofacial dysmorphism, pigmentary skin anomalies, epilepsy, and a variety of congenital malformations. Prenatally, PKS can be suspected in the presence of ultrasound anomalies: diaphragmatic hernia, rhizomelic micromelia, hydrops fetalis, fetal overweight, ventriculomegaly in the central nervous system, congenital heart defects, or absent visualization of the stomach. In all these cases, a detailed genetic study is required. PKS is diagnosed by prenatal genetic analysis through chorionic villus sampling, genetic amniocentesis, and cordocentesis. CASE REPORT: We report two cases of PKS with prenatal diagnosis of isochromosome 12p made by cytogenetic studies. The first case is of a 36-year-old pregnant woman who underwent genetic chorionic villus sampling at 13th weeks of gestation after 1st trimester prenatal ultrasound revealed clinical features of PKS: flat nasal bridge and fetal hydrops. The second case is of a 32-year-old pregnant woman with genetic amniocentesis at 17th weeks of gestation that showed mos46,XX[21]/47,XX,+i(12p) associated to PKS. CONCLUSION: New molecular cytogenetic techniques array comparative genomic hybridization and fluorescence in-situ hybridization in association with conventional karyotype are pivotal innovative tools to search for chromosomic anomalies and for a complete prenatal diagnosis, especially in cases such as PKS where array comparative genomic hybridization analysis alone could not show mosaicism of i(12p).

New Metacentric Populations and Phylogenetic Hypotheses Involving Whole-Arm Reciprocal Translocation in Mus musculus domesticus from Sicily, Southern Italy.

The house mouse Mus musculus domesticus is characterized by more than 100 metacentric populations, due to the occurrence of Robertsonian (Rb) fusions, together with the standard all-telocentric karyotype (2n = 40). We examined G-banded karyotypes of 18 mice from 10 localities in Sicily and describe 3 new metacentric populations: 'Ragusa Ibla' (IRAG), 2n = 33-36, Rb(2.4), Rb(5.6), Rb(9.16), Rb(13.17); 'Piana degli Albanesi' (IPIA), 2n = 23, Rb(1.18), Rb(2.15), Rb(3.5), Rb(4.12), Rb(6.11), Rb(7.8), Rb(9.16), Rb(10.14), Rb(13.17); 'Trapani' (ITRA), 2n = 22, Rb(1.18), Rb(2.15), Rb(3.7), Rb(4.12), Rb(5.9), Rb(6.11), Rb(8.16), Rb(10.14), Rb(13.17). Three mice belonged to the previously reported 'Castelbuono' race (ICAS), 2n = 24, which is very similar to the nearby 'Palermo' (IPAL) race, 2n = 26. Three Rb fusions not yet observed in wild mouse populations were identified: Rb(3.5), Rb(3.7) and Rb(5.9). Rb fusions shared among 4 races (IPIA, IRAG, ICAS, and IPAL) allowed us to describe their potential phylogenetic relationships. We obtained 2 alternative phylogenetic trees. The differences between them are mainly due to various modes of formation of IPIA and ITRA. In the first hypothesis, the specific Rb fusions occurred independently. In the second, those of IRAG originated from those of IPIA via whole-arm reciprocal translocations.

A new case of interstitial 1q 25.3-32.1 deletion: cytogenetic analysis molecular characterization and ultrasound findings.

INTRODUCTION: deletion of long arm of chromosome 1(1q-) is a rare condition. Clinical features include Dwarfism, severe mental retardation, microcephaly and short neck delineating the "intermediate 1q deletion syndrome". CASE REPORT: we report a new case of interstitial deletion of the long arm of chromosome 1, diagnosed in a 22+3 weeks gestation fetus in which cytogenetic analysis localized a loss of genetic materials of 18Mb in the 1q25.3-32.1. Fetal ultrasound showed neurodegenerative defects resembling Dandy-Walker's syndrome and bilateral clubfoot. CONCLUSIONS: clinical characteristics of our case are markedly mild. This suggests that the type and the extension of the mutation obtained through cytogenetic studies, CGH array and ultrasound evaluation should be taken into account for prognostic evaluation and management of these patients.

Six consecutive false positive cases from cell-free fetal DNA testing in a single referring centre.

INTRODUCTION: recent studies have proposed the introduction of cell-free fetal DNA testing (NIPT-Non Invasive Prenatal Testing) in routine clinical practice emphasizing its high sensibility and specificity. In any case, false positive and false negative findings may result from placental mosaicism, because cell-free fetal DNA originates mainly from placenta. CASE: WE REPORT SIX CASES OF WOMEN WHO UNDERWENT CHORIONIC VILLUS SAMPLING (CVS) OR AMNIOCENTESIS TO CONFIRM THE RESULTS FROM NIPT: two Turner syndromes, two Triple X, one Patau syndrome, one Edward syndrome. RESULTS: using classic cytogenetic analysis and, also, Array - Comparative Genomic Hybridization (Array CGH) the karyotype of all 5 fetuses was found to be normal. CONCLUSION: results from NIPT must always be confirmed by invasive prenatal diagnosis. It is mandatory to inform the patient that the CVS and amniocentesis still represent the only form of prenatal diagnostic test available.

Retrospective study evaluating the performance of a first-trimester combined screening for trisomy 21 in an Italian unselected population.

OBJECTIVES: to assess the performance of a combined first-trimester screening for trisomy 21 in an unselected Italian population referred to a specialized private center for prenatal medicine. METHODS: a retrospective validation of first-trimester screening algorithms [risk calculation based on maternal age and nuchal translucency (NT) alone, maternal age and serum parameters (free ß-hCG and PAPP-A) alone and a combination of both] for fetal aneuploidies evaluated in an unselected Italian population at Artemisia Fetal-Maternal Medical Centre in Rome. All measurements were performed between 11(+0) and 13(+6) weeks of gestation, between April 2007 and December 2008. RESULTS: of 3,610 single fetuses included in the study, we had a complete follow-up on 2,984. Fourteen of 17 cases of trisomy 21 were detected when a cut-off of 1:300 was applied [detection rate (DR) 82.4%, 95% confidence interval (CI) 64.2-100; false-positive rate (FPR) 4.7%, 95% CI 3.9-5.4; false-negative rate (FNR) 17.6%, 95% CI 0-35.8%]. CONCLUSION: in our study population the detection rate for trisomy 21, using the combined risk calculation based on maternal age, fetal NT, maternal PAPP-A and free ß-hCG levels, was superior to the application of either parameter alone. The algorithm has been validated for first trimester screening in the Italian population.

Discordance between karyotype from amniotic fluid and postnatal lymphocyte cultures.

Discordance between karyotype seen from amniocentesis and from neonatal blood is a very unusual condition with different possible causes.We present a case of discordance between prenatal cytogenetic diagnosis from amniotic fluid and post-natal cytogenetic diagnosis from lymphocyte cultures.

Prenatal Diagnosis of a Fetus with de novo Supernumerary Ring Chromosome 16 Characterized by Array Comparative Genomic Hybridization.

A fetus with de novo ring chromosome 16 is presented. At 20 weeks' gestation, ultrasound examination demonstrated bilateral clubfoot, bilateral renal pyelectasis, hypoplasia of the corpus callosum, and transposition of the great vessel. Amniocentesis was performed. Chromosome analysis identified a ring chromosome 16 [47,XY,r(16)] and array comparative genomic hybridization (a-CGH) demonstrated that the ring included the euchromatic portion 16p11.2. Postmortem examination confirmed prenatal findings. This is the first case of de novo ring chromosome 16 diagnosed prenatally with a new phenotypic pattern and also reinforces the importance of offering amniocentesis with a-CGH if fetal anomalies are detected.

The use of DHPLC (Denaturing High Performance Liquid Chromatography) in II level screening of the CFTR gene in Prenatal Diagnosis.

OBJECTIVE: The aim of the study is to evaluate the role of Denaturing High Performance Liquid Chromatography (DHPLC) in the second level screening of the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) gene. METHODS: A 9-month prospective study, between June 2008 and March 2009 at Artemisia Fetal Medical Centre, included 3829 samples of amniotic fluid collected from women undergoing mid-trimester amniocentesis.The genetic diagnosis of CF was based on research of the main mutations of the CFTR gene on fetal DNA extracted from the amniocytes, (first level screening) using different commercial diagnostic systems. A second level screening using DHPLC, on the amniotic fluid and on a blood sample from the couple, was offered in case of fetuses heterozygous at first level screening. RESULTS: Of 3829 fetuses, 134 were found to be positive, 129 heterozygous and 5 affected. Of the 129 couples, following appropriate genetic counselling, 53 requested a second level screening. Through the use of DHPLC, 44 couples were found to be negative, and in nine couples, nine rare mutations were identified. CONCLUSIONS: The first level screening can be useful to evidence up to 75% of the CF mutations. The second level screening can identify a further 10% of mutant alleles. DHPLC was found to be a reliable and specific method for the rapid identification of the rare CFTR mutations which were not revealed in initial first level screening.

Antibiotic prophylaxis before second-trimester genetic amniocentesis (APGA): a single-centre open randomised controlled trial.

OBJECTIVE: To compare procedure-related pregnancy loss after second-trimester genetic amniocentesis in women given an antibiotic prophylaxis and controls. METHODS: Prospective, open randomised controlled single-centre study between January 1999 and December 2005 at Artemisia Fetal Maternal Medical Centre. A follow-up within 4 weeks after the procedure was done.Of 36,347 eligible women, 1424 refused to participate and 34,923 were enrolled and randomised with unequal chance of selection, 21,991 were assigned to treatment group and 12,932 were assigned to the control group, and did not receive any placebo. Oral azithromycin, 500 mg per day, was administered 3 days before amniocentesis. The primary endpoint was the procedure-related pregnancy loss. The secondary endpoint was the rate of preterm premature rupture of membranes. RESULTS: The rate of abortion related to the amniocentesis was 7/21 219 women (0.03%, 95% CI 0.009-0.057) in the intervention group, and 36/12 529 (0.28%, 0.28-0.30) in controls (p = 0.0019). The rate of preterm premature rupture of membranes was 14/21 219 (0.06%, 0.031-0.101) in the intervention group, and 140/12 529 (1.12%, 0.94-1.30) in the control group (p = 0.001). CONCLUSIONS: Antibiotic prophylaxis before second-trimester amniocentesis reduced the risk of abortion and of rupture of the membranes.

Prenatal screening of Cystic Fibrosis: a single centre experience.

OBJECTIVE: The gene responsible for the pathogenesis of cystic fibrosis has been known for over 15 years and represent the most common autosomal recessive disease in the european population. We aimed to investigate the incidence of this condition during fetal life. METHODS: In the past 10 years we examined in our centre 25393 fetuses of women underwent to amniocentesis. We carried out the examination of the most frequent mutations which enable, according to the literature data, the identification of almost 80% of the affected alleles. RESULT: We identified 922 heterozygous and 9 homozygous for the mutation. The frequency of heterozygousin the examined sample was 1/27,5 while that of the affected was 1/2821. CONCLUSION: We encourage new thoughts regarding the diagnostic validity of the most frequent panel of mutations among the italian population in order to exclude never encountered mutations and the insertion of other more significant mutations.

A case report of a meiotic segregation study on a small supernumerary marker chromosome.

Small supernumerary marker chromosomes (sSMCs) have been described from all human chromosomes with different sizes and shapes. However, it is difficult to know the clinical manifestations associated with them, because such knowledge depends on the size, presence of euchromatic material, degree of mosaicism and/or uniparental disomy (UPD).A case report of a familial small supernumerary marker chromosome (sSMC) through a structural and a segregation study is reported.