Emmanouil Manolakos

High frequency of the TARDBP p.Ala382Thr mutation in Sardinian patients with amyotrophic lateral sclerosis

Orrù S, Manolakos E, Orrù N, Kokotas H, Mascia V, Carcassi C, Petersen MB. High frequency of the TARDBP p.Ala382Thr mutation in Sardinian patients with amyotrophic lateral sclerosis.

Testis development in the absence of SRY: chromosomal rearrangements at SOX9 and SOX3

Duplications in the ~2 Mb desert region upstream of SOX9 at 17q24.3 may result in familial 46,XX disorders of sex development (DSD) without any effects on the XY background. A balanced translocation with its breakpoint falling within the same region has also been described in one XX DSD subject. We analyzed, by conventional and molecular cytogenetics, 19 novel SRY-negative unrelated 46,XX subjects both familial and sporadic, with isolated DSD. One of them had a de novo reciprocal t(11;17) translocation. Two cases carried partially overlapping 17q24.3 duplications ~500 kb upstream of SOX9, both inherited from their normal fathers. Breakpoints cloning showed that both duplications were in tandem, whereas the 17q in the reciprocal translocation was broken at ~800 kb upstream of SOX9, which is not only close to a previously described 46,XX DSD translocation, but also to translocations without any effects on the gonadal development. A further XX male, ascertained because of intellectual disability, carried a de novo cryptic duplication at Xq27.1, involving SOX3. CNVs involving SOX3 or its flanking regions have been reported in four XX DSD subjects. Collectively in our cohort of 19 novel cases of SRY-negative 46,XX DSD, the duplications upstream of SOX9 account for ~10.5% of the cases, and are responsible for the disease phenotype, even when inherited from a normal father. Translocations interrupting this region may also affect the gonadal development, possibly depending on the chromatin context of the recipient chromosome. SOX3 duplications may substitute SRY in some XX subjects.

The use of array-CGH in a cohort of Greek children with developmental delay

BackgroundThe genetic diagnosis of mental retardation (MR) is difficult to establish and at present many cases remain undiagnosed and unexplained. Standard karyotyping has been used as one of the routine techniques for the last decades. The implementation of Array Comparative Genomic Hybridization (array-CGH) has enabled the analysis of copy number variants (CNVs) with high resolution. Major cohort studies attribute 11% of patients with unexplained mental retardation to clinically significant CNVs. Here we report the use of array-CGH for the first time in a Greek cohort. A total of 82 children of Greek origin with mean age 4.9 years were analysed in the present study. Patients with visible cytogenetic abnormalities ascertained by standard karyotyping as well as those with subtelomeric abnormalities determined by Multiplex Ligation-dependent Probe Amplification (MLPA) or subtelomeric FISH had been excluded.ResultsFourteen CNVs were detected in the studied patients. In nine patients (11%) the chromosomal aberrations were inherited from one of the parents. One patients showed two duplications, a 550 kb duplication in 3p14.1 inherited from the father and a ~1.1 Mb duplication in (22)(q13.1q13.2) inherited from the mother. Although both parents were phenotypically normal, it cannot be excluded that the dual duplication is causative for the patients clinical profile including dysmorphic features and severe developmental delay. Furthermore, three de novo clinically significant CNVs were detected (3.7%). There was a ~6 Mb triplication of 18q21.1 in a girl 5 years of age with moderate MR and mild dysmorphic features and a ~4.8 Mb duplication at (10)(q11.1q11.21) in a 2 years old boy with severe MR, multiple congenital anomalies, severe central hypotonia, and ataxia. Finally, in a 3 year-old girl with microcephaly and severe hypotonia a deletion in (2)(q31.2q31.3) of about ~3.9 Mb was discovered. All CNVs were confirmed by Fluorescence in situ hybridization (FISH). For the remaining 9 patients the detected CNVs (inherited duplications or deletions of 80 kb to 800 kb in size) were probably not associated with the clinical findings.ConclusionsGenomic microarrays have within the recent years proven to be a highly useful tool in the investigation of unexplained MR. The cohorts reported so far agree on an around 11% diagnostic yield of clinically significant CNVs in patients with unexplained MR. Various publicly available databases have been created for the interpretation of identified CNVs and parents are analyzed in case a rare CNV is identified in the child. We have conducted a study of Greek patients with unexplained MR and confirmed the high diagnostic value of the previous studies. It is important that the technique becomes available also in less developed countries when the cost of consumables will be reduced.

Detection of congenital heart defects throughout pregnancy; impact of first trimester ultrasound screening for cardiac abnormalities.

Objective: To evaluate prospectively the efficacy to screen for congenital heart defects (CHD) during the first trimester nuchal translucency (NT) ultrasound examination by assessing the four chambers’ view of fetal heart. Methods: Pregnancies that were examined prospectively by ultrasound in the first trimester (11th–14th week), the second (19th–24th week) and third trimester were included in the study. 3774 fetuses were examined and fetal heart was assessed during the NT scan by examining the four chambers view. Detailed echocardiography was performed during the anomaly and growth scans. Diagnosis of congenital heart defects (CHD) was further confirmed by a fetal cardiologist. Results: The four chambers view was obtained in 99.52% of the cases. CHD were diagnosed in 29 fetuses (0.77%). Thirteen cases (44.8%) were detected during the 11–13 weeks’ scan, 14 cases (48.3%) during the anomaly scan, 1 CHD (3.5%) during the third trimester scan and 1 case (3.5%) postpartum. Conclusion: Assessment of the four chambers of fetal heart early in pregnancy was feasible and allowed the detection of 45% of CHD. Additional parameters of fetal cardiac anatomy during the NT scan may further improve the detection rate providing pregnancy management information early in the first trimester.

Dual testing with QF-PCR and karyotype analysis for prenatal diagnosis of chromosomal abnormalities. Evaluation of 13,500 cases with consideration of using QF-PCR as a stand-alone test according to referral indications.

Evaluate the results obtained from Quantitative Fluorescent (QF)‐PCR and conventional karyotype analysis to determine the advantages and disadvantages of dual testing in prenatal diagnosis.

Routine use of array comparative genomic hybridization (aCGH) as standard approach for prenatal diagnosis of chromosomal abnormalities. Clinical experience of 1763 prenatal cases.

This study aims to evaluate the diagnostic yield of comparative genomic hybridization microarrays (aCGH) and compare it with conventional karyotype analysis of standard >5‐Mb resolution.

A patient with partial trisomy 21 and 7q deletion expresses mild Down syndrome phenotype

BACKGROUND Down syndrome (DS) is the most common aneuploidy in live-born individuals and it is well recognized with various phenotypic expressions. Although an extra chromosome 21 is the genetic cause for DS, specific phenotypic features may result from the duplication of smaller regions of the chromosome and more studies need to define genotypic and phenotypic correlations. CASE REPORT We report on a 26 year old male with partial trisomy 21 presenting mild clinical symptoms relative to DS including borderline intellectual disability. In particular, the face and the presence of hypotonia and keratoconus were suggestive for the DS although the condition remained unnoticed until his adult age array comparative genomic hybridization (aCGH) revealed a 10.1 Mb duplication in 21q22.13q22.3 and a small deletion of 2.2 Mb on chromosomal band 7q36 arising from a paternal translocation t(7;21). The 21q duplication encompasses the gene DYRK1. CONCLUSION Our data support the evidence of specific regions on distal 21q whose duplication results in phenotypes recalling the typical DS face. Although the duplication region contains DYRK1, which has previously been implicated in the causation of DS, our patient has a borderline IQ confirming that their duplication is not sufficient to cause the full DS phenotype.

Prenatal detection of full monosomy 21 in a fetus with increased nuchal translucency: Molecular cytogenetic analysis and review of the literature

Full monosomy 21 is an extremely rare chromosomal disorder. A 38‐year‐old woman attended a first trimester scan. Ultrasound (U/S) imaging of the fetus at 12 weeks of gestation showed features of increased nuchal translucency measurement (12 mm). Chorionic villi sampling (CVS) was performed after genetic counseling. At 16 weeks of gestation the fetus showed U/S characteristics of severe intrauterine growth restriction, generalized edema and hydrothorax. Cytogenetic examination was performed using quantitative fluorescent polymerase chain reaction analysis, standard Giesma banding and fluorescent in situ hybridization analysis. Non‐mosaic full monosomy 21 was detected and the parents opted to terminate the pregnancy. Microsatellite analysis demonstrated maternal origin of the single chromosome. This case represents one of the few cases of prenatally diagnosed full monosomy 21 confirmed only by CVS, in which the parental origin of the single chromosome was determined.

Characterization of a prenatally assessed de novo supernumerary minute ring chromosome 20 in a phenotypically normal male

BackgroundThe heterogeneous group of small supernumerary marker chromosomes (sSMCs) presents serious counseling problems, especially if they are present de novo and diagnosed prenatally. The incidence has been estimated at 1 in 1000 prenatal samples. We present a case of mosaic sSMC diagnosed prenatally after amniocentesis. The sSMC was characterized by various molecular cytogenetic techniques and determined to be a r(20) chromosome. After genetic counseling, the parents decided to continue the pregnancy, and a boy with minor phenotypic variants was born after 39 weeks of pregnancy. The case is compared with four other cases of prenatally detected r(20) mosaicism.ResultsHere we describe a 3 months old male child with normal pre- and postnatal development and with a de novo ring supernumerary marker chromosome in amniocytes cultures. Using new fluorescence in situ hybridization (FISH) techniques, three distinguishable sSMCs (cryptic mosaicism), all derived from chromosome 20, were observed, including ring and minute chromosomes. This heterogeneity was impossible to detect by the conventional G-banding technique or conventional FISH technique that were used before the application of new FISH techniques (subcentromere-specific multicolor-FISH [subcenM-FISH]) and a probe, specific for the 20p12.2 band. The sSMC present in 25% of the cells was present as r(20)(::p12.2~12.3->q11.1::)[5]/r(20;20)(::p12.1->q11.1::q11.1 >p12.1::)[2]/min(20;20)(:p12.1->q11.1::q11.1->p12.1:)[1]. The final karyotype was 47,XY,+r(20)[25%]/46,XY[75%].ConclusionWe emphasize the importance of application of molecular cytogenetics in a prenatally diagnostic laboratory and description of more cases to enable a better genetic counseling and risk evaluation.

Prenatal diagnosis of Wolf-Hirschhorn syndrome confirmed by comparative genomic hybridization array: report of two cases and review of the literature

Wolf-Hirschhorn syndrome (WHS) is a well known genetic condition caused by a partial deletion of the short arm of chromosome 4. The great variability in the extent of the 4p deletion and the possible contribution of additional genetic rearrangements lead to a wide spectrum of clinical manifestations. The majority of the reports of prenatally diagnosed WHS cases are associated with large 4p deletions identified by conventional chromosome analysis; however, the widespread clinical use of novel molecular techniques such as array comparative genomic hybridization (a-CGH) has increased the detection rate of submicroscopic chromosomal aberrations associated with WHS phenotype. We provide a report of two fetuses with WHS presenting with intrauterine growth restriction as an isolated finding or combined with oligohydramnios and abnormal Doppler waveform in umbilical artery and uterine arteries. Standard karyotyping demonstrated a deletion on chromosome 4 in both cases [del(4)(p15.33) and del(4)(p15.31), respectively] and further application of a-CGH confirmed the diagnosis and offered a precise characterization of the genetic defect. A detailed review of the currently available literature on the prenatal diagnostic approach of WHS in terms of fetal sonographic assessment and molecular cytogenetic investigation is also provided.