Prenatal diagnosis of chromosomal abnormalities in fetuses with abnormal cardiac ultrasound findings: evaluation of chromosomal microarray-based analysis.

OBJECTIVES: To assess the frequency of karyotype abnormalities and chromosome 22q11.2 deletion syndrome among fetuses with abnormal cardiac ultrasound findings, and to evaluate the clinical value of chromosomal microarray-based analysis (CMA) in the study of such pregnancies. METHODS: First, we carried out retrospective analysis of karyotype abnormalities and 22q11.2 deletion syndrome cases diagnosed between January 2009 and December 2011 in our center among fetuses with abnormal cardiac ultrasound findings (n = 276). Second, CMA was performed in 51 of the fetuses with such findings, normal karyotype and negative or no 22q11.2 deletion syndrome study, and in the only fetus with a heart defect and an apparently balanced de novo chromosomal rearrangement. RESULTS: Out of the 276 pregnancies with abnormal cardiac ultrasound findings, karyotyping revealed a chromosomal abnormality in 44 (15.9%). Of fetuses with normal karyotype in which 22q11.2 deletion syndrome studies were performed, 6.4% (5/78) had this microdeletion syndrome. Among fetuses with abnormal cardiac findings, normal karyotype and negative or no 22q11.2 deletion syndrome study that underwent CMA, the detection rate of pathogenic copy number variants not detected by conventional cytogenetics was 2.0% (1/51), and no variants of uncertain clinical significance were found. In the fetus with a heart defect and an apparently balanced de novo chromosomal rearrangement, CMA revealed that the rearrangement was not truly balanced. CONCLUSIONS: In the assessment of genetic abnormalities in pregnancies with abnormal cardiac ultrasound findings, the diagnostic yield may be increased by 2% if CMA is used as a complementary tool to conventional cytogenetics. Our results suggest that CMA could be a good alternative to karyotyping in these pregnancies.

El diagnóstico citogenético prenatal aquí y ahora. Validez actual y futura del cariotipo / The cytogenetic prenatal diagnosis nowadays. Present and future value of karyotype

El cariotipo convencional obtenido mediante las técnicasde bandas es considerado como el gold standard deldiagnóstico prenatal citogenético; nos da la información detodos los cromosomas, es decir de todo el genoma, con un nivelde resolución limitado pero aceptable. Está incorporado en lapráctica asistencial de la mayoría de hospitales de tercer nivelde nuestro país, así como en centros sanitarios y laboratoriosprivados. En buena parte de ellos se aplican además otrastécnicas complementarias de citogenética molecular paracompletar algún diagnóstico concreto.Durante estos últimos años se han ido desarrollando técnicasmoleculares que permiten superar determinados inconvenientesdel cultivo celular y análisis citogenético, como laduración del cultivo (Quantitative Fluorescent PolymeraseChain Reaction [QF-PCR]) o el limitado poder de resolución delos cromosomas bandeados (técnicas de Array Genomic Hybridization[AGH]). Sin embargo, estas técnicas también tienensus desventajas con respecto al cariotipo: resultados parciales,coste económico elevado, falta de información de las reorganizacionescromosómicas (fundamental para el consejo genéticode la gestante y su familia), y problemas de interpretaciónde los resultados debido a la existencia de Copy Number Variants(CNV) polimórficas o benignas, con el agravante de contarsolamente con el examen ecográfico del «paciente».A corto plazo, por su relación coste-beneficio, el futurodel cariotipo como técnica básica de diagnóstico prenatalcitogenético parece asegurado. El empleo de técnicas complementariasmoleculares o citogenético-moleculares, paracompletar los análisis en los casos que lo requieran, va a sercada vez más necesario...(AU)

The conventional karyotype, obtained using bandingtechniques, is considered the «gold standard» in prenatal cytogeneticdiagnosis: it gives genome-wide information (from allthe chromosomes), with limited but acceptable resolution.Most of the great public hospitals of our country, as well asmany private sanitary centres and laboratories, offer conventionalkaryotype as routine prenatal diagnosis, and in most ofthem, complementary molecular-cytogenetic techniques arealso available for application in special cases.New molecular techniques have been developed duringthe last years, which overcome certain drawbacks of cellculture and cytogenetic analysis, such as the duration of theculture (Quantitative Fluorescent Polymerase Chain Reaction[QF-PCR]) or the limited resolution of the banded chromosomes(Array Genomic Hybridization [AGH] techniques).However, these techniques also have disadvantages with respectto the karyotype: partial results, high costs, lack of informationabout chromosome rearrangements (essential forthe genetic counselling to the pregnant woman and herfamily), and interpretation problems of results due to theexistence of benign Copy Number Variants (CNV) and difficultiesto correlate the findings with the fetal phenotype,only visible through echography.In the near future, the karyotype will remain as the basictool in cytogenetic prenatal diagnosis, due to its costbenefitrelation. The use of complementary molecular ormolecular-cytogenetic techniques in special cases will becomemore and more frequent.In the distant future, with the rising technological advances,the karyotype will probably become the complementary technique, but it will be also necessary in most casesfor validation (Fluorescence in Situ Hybridization [FISH])and, above all, as an essential tool for genetic counselling(AU)

Age and origin of major Smith-Lemli-Opitz syndrome (SLOS) mutations in European populations.

BACKGROUND: Smith-Lemli-Opitz syndrome (SLOS) (MIM 270 400) is an autosomal recessive multiple congenital anomalies/mental retardation syndrome caused by mutations in the Delta7-sterol reductase (DHCR7, E.C.1.3.1.21) gene. The prevalence of SLOS has been estimated to range between 1:15000 and 1:60000 in populations of European origin. METHODS AND RESULTS: We have analysed the frequency, origin, and age of DHCR7 mutations in European populations. In 263 SLOS patients 10 common alleles (c.964-1G>C, p.Trp151X, p.Thr93Met, p.Val326Leu, p.Arg352Trp, p.Arg404Cys, p.Phe302Leu, p.Leu157Pro, p.Gly410Ser, p.Arg445Gln) were found to constitute approximately 80% of disease-causing mutations. As reported before, the mutational spectra differed significantly between populations, and frequency peaks of common mutations were observed in North-West (c.964-1G>C), North-East (p.Trp151X, p.Val326Leu) and Southern Europe (p.Thr93Met). SLOS was virtually absent from Finland. The analysis of nearly 8000 alleles from 10 different European populations confirmed a geographical distribution of DHCR7 mutations as reported in previous studies. The common Null mutations in Northern Europe (combined ca. 1:70) occurred at a much higher frequency than expected from the reported prevalence of SLOS. In contrast the most common mutation in Mediterranean SLOS patients (p.Thr93Met) had a low population frequency. Haplotypes were constructed for SLOS chromosomes, and for wild-type chromosomes of African and European origins using eight cSNPs in the DHCR7 gene. The DHCR7 orthologue was sequenced in eight chimpanzees (Pan troglodytes) and three microsatellites were analysed in 50 of the SLOS families in order to estimate the age of the three major SLOS-causing mutations. CONCLUSIONS: The results indicate a time of first appearance of c.964-1G>C and p.Trp151X some 3000 years ago in North-West and North-East Europe, respectively. The p.Thr93Met mutations on the J haplotype has probably first arisen approximately 6000 years ago in the Eastern Mediterranean. Together, it appears that a combination of founder effects, recurrent mutations, and drift have shaped the present frequency distribution of DHCR7 mutations in Europe.

Identification of 14 novel mutations in DHCR7 causing the Smith-Lemli-Opitz syndrome and delineation of the DHCR7 mutational spectra in Spain and Italy.

The Smith-Lemli-Opitz syndrome (SLOS) is a phenotypically variable metabolic malformation and mental retardation syndrome for which more than 80 mutations in the DHCR7 disease-causing gene have been described. The DHCR7 mutational spectra differ significantly in different areas of Europe, and several common putative founder mutations account for a substantial fraction of all mutations in some ethnic groups. Here we have analysed 47 SLOS patients and describe 14 newly identified mutations in 18 SLOS patients of Ashkenazi Jewish, Austrian, British, German, Italian, Irish, Polish, Portuguese, and Spanish origins. Half of the new mutations are in the transmembrane domains of the protein. In addition, there were two null mutations, one mutation in the 4th cytoplasmic loop, two mutations in the first and last codons, and three mutations in other regions such as the second cytoplasmic loop and the first endoplasmic loop. The analysis included 20 Spanish and 12 Italian SLOS patients and revealed very different mutation spectra in these patients compared to previously described patients from Czechoslovakia, Germany, Poland, and the UK and implicated p.Thr93Met on the J haplotype as the most frequent Mediterranean founder mutation.

FISH on sperm heads allows the analysis of chromosome segregation and interchromosomal effects in carriers of structural rearrangements: results in a translocation carrier, t(5;8)(q33;q13).

Using three-color fluorescence in situ hybridization (FISH) with specific DNA probes, we have determined the chromosome segregation pattern of the translocated chromosomes in a human male carrier of a reciprocal translocation, t(5;8)(q33;q13). At the same time, we have assessed the possible interchromosomal effect on pair 21 using dual-color FISH. The segregation results showed that a 45.12% of the spermatozoa analyzed resulted from alternate segregation, 38.31% resulted from adjacent I, 6.97% from adjacent II, and 6.56% from 3:1 segregation. Finally, 1.23% could be either diploid sperm or 4:0 segregation. In both types of adjacent segregations, an excess of products containing short translocated segments (adjacent I) and interstitial regions (adjacent II) were found. Products resulting from the presence of an interstitial chiasma in pair 5 (1.26%), were found much more frequently (P < 0.0001) than those resulting from an interstitial chiasma in pair 8 (0.13%) (evaluated after adjacent II segregation). In 3:1 segregation, the products containing one chromosome were observed more frequently than those containing three chromosomes (P < 0.0001). No evidence of an interchromosomal effect on chromosome 21 was detected, the percentage of disomy 21 being similar to that in the controls (0.23% vs. 0.37%). However, the percentage of diploid sperm (1.18%) was significantly higher (P < 0. 0001) than that in the controls (0.27%). FISH therefore appears to be a useful technique for assessing the percentage of abnormal sperm in translocation carriers. Their application in assisted reproduction centers could offer patients more accurate genetic counseling.

[Distal trisomy 14q caused by a paternal translocation (6;14)]. / Trisomía 14q distal debida a una translocación paterna (6;14).

A female child was found to have a partial duplication of the long arm of chromosome 14 (14q24----qter). This duplication was due to paternal balanced translocation 46,XY,t(6;14) (q27;q24). Patient had craniofacial dysmorphism with microphtalmia, flat nasal bridge, lowest malformed ears, short neck, dolicocolon and mental retardation. A comparison is made between other reported cases in the literature and this observation. Most of the anomalies described in these published reports are not shared by the present patient. This can be explained by the simultaneous presence or not of a partial deletion and the size of the duplicated segment.

De novo interstitial deletion of the long arm of chromosome 2 in a malformed newborn with a karyotype: 46,XY,del(2)(q12q14).

A male neonate with severe malformations, including facial dysmorphism, a short neck, postaxial-hexadactyly of the toes, congenital heart disease, hydronephrosis, imperforate anus and agenesis of corpus callosum, is described. His karyotype was 46,XYdel(2)(q12q14).