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Background Congenital malformations involving the Mullerian ducts are observed in around 5% of infertile women. Complete aplasia of the uterus, cervix, and upper vagina, also termed Mullerian aplasia or Mayer–Rokitansky–Kuster–Hauser (MRKH) syndrome, occurs with an incidence of around 1 in 4500 female births, and occurs in both isolated and syndromic forms. Previous reports have suggested that a proportion of cases, especially syndromic cases, are caused by variation in copy number at different genomic loci. Methods In order to obtain an overview of the contribution of copy number variation to both isolated and syndromic forms of Mullerian aplasia, copy number assays were performed in a series of 63 cases, of which 25 were syndromic and 38 isolated. Results A high incidence (9/63, 14%) of recurrent copy number variants in this cohort is reported here. These comprised four cases of microdeletion at 16p11.2, an autism susceptibility locus not previously associated with Mullerian aplasia, four cases of microdeletion at 17q12, and one case of a distal 22q11.2 microdeletion. Microdeletions at 16p11.2 and 17q12 were found in 4/38 (10.5%) cases with isolated Mullerian aplasia, and at 16p11.2, 17q12 and 22q11.2 (distal) in 5/25 cases (20%) with syndromic Mullerian aplasia. Conclusion The finding of microdeletion at 16p11.2 in 2/38 (5%) of isolated and 2/25 (8%) of syndromic cases suggests a significant contribution of this copy number variant alone to the pathogenesis of Mullerian aplasia. Overall, the high incidence of recurrent copy number variants in all forms of Mullerian aplasia has implications for the understanding of the aetiopathogenesis of the condition, and for genetic counselling in families affected by it.

We report a 35-year-old male with a ring chromosome 12 originally diagnosed 20 years prior to presentation with an ischemic stroke. Array CGH analysis revealed a sub-microscopic microdeletion and microduplication within 12p13.3 and a microdeletion in 12q24.33. FISH analysis further revealed that the duplication was in an inverted orientation and included exons 35-52 of the dosage-sensitive Von Willebrand Factor (VWF) gene. Partial duplication of this gene, which has a role in the clotting cascade, suggests a potential mechanism for generating a pro-thrombotic state that may have contributed to a premature cerebrovascular event. Evidence of raised VWF antigen levels and VWF activity levels in the highest quartile provides support for this hypothesis. This case illustrates that when a ring chromosome is identified, the possibility of cryptic genomic rearrangements needs to be considered as these may have implications in predicting natural history.

Copy number variants and indels in 251 families with evidence of X-linked intellectual disability (XLID) were investigated by array comparative genomic hybridization on a high-density oligonucleotide X chromosome array platform. We identified pathogenic copy number variants in 10% of families, with mutations ranging from 2 kb to 11 Mb in size. The challenge of assessing causality was facilitated by prior knowledge of XLID-associated genes and the ability to test for cosegregation of variants with disease through extended pedigrees. Fine-scale analysis of rare variants in XLID families leads us to propose four additional genes, PTCHD1, WDR13, FAAH2, and GSPT2, as candidates for XLID causation and the identification of further deletions and duplications affecting X chromosome genes but without apparent disease consequences. Breakpoints of pathogenic variants were characterized to provide insight into the underlying mutational mechanisms and indicated a predominance of mitotic rather than meiotic events. By effectively bridging the gap between karyotype-level investigations and X chromosome exon resequencing, this study informs discussion of alternative mutational mechanisms, such as noncoding variants and non-X-linked disease, which might explain the shortfall of mutation yield in the well-characterized International Genetics of Learning Disability (IGOLD) cohort, where currently disease remains unexplained in two-thirds of families.

Cytogenetically visible imbalances without phenotypic effect are still rare despite the extent of large-scale copy number variation in the normal population revealed by array CGH. Here we report on a phenotypically normal 30-year-old female with a de novo, cytogenetically visible, interstitial deletion of band 4q34. She was referred following three successive miscarriages, one of which was an intra-uterine death with subendocardial fibroelastosis and dilated cardiomyopathy. There was no other notable medical or family history, she was of normal intelligence and had no dysmorphic features. FISH and Array CGH with a customized 1 Mb BAC array showed that the deletion is a minimum of 9.3 and a maximum of 10.7 Mb in size, between approximately 173 Mb in 4q34.1 and approximately 182 Mb in 4q34.3. The deletion contains only 23 known coding genes giving a low average gene density of approximately 2 genes/Mb. This case further illustrates that (1) sizeable imbalances can be associated with apparent phenotypic normality, (2) gene density is a better guide to possible phenotypic consequences than aberration size, and (3) it is not always safe to assume that de novo imbalances will be causal.

Background: Microdeletions at 17q21.31 have recently been shown to cause a novel syndrome. Here we identify the reciprocal 17q21.31 duplication syndrome in 4 patients. Method: Patients with the 17q21.31 duplication were identified by screening a large cohort of patients (n = 13 070) with mental retardation and congenital malformation by comparative genomic hybridisation microarray. Parental origin was investigated in 3 patients by quantitative polymerase chain reaction and microsatellite genotyping. Results: In three cases it was possible to show that duplication arose de novo. Intellectual skills range from normal to mild mental retardation. Patients are characterised by poor social interaction, with relationship difficulties, reminiscent of autistic spectrum disorders. Other features are rather variable with no striking common phenotypic features. Parental origin was investigated for 3 patients. In all cases duplication was of maternal origin either through interchromosomal (2 cases) or interchromatid (1 case) rearrangement. The 3 mothers are all carriers of the inverted H2 haplotype, emphasising the role of local genomic architecture alteration as a predisposing factor for this duplication. Conclusion: Autistic features observed in our patients suggest that genes in the duplicated interval should be considered as candidates for disorders in the autistic spectrum. Other phenotypic observations are rather variable or aspecific. This adds 17q21.31 duplications to a growing group of recently identified genomic disorders with variable penetrance and expressivity.

Molecular karyotyping is being increasingly applied to delineate novel disease causing microaberrations and related syndromes in patients with mental retardation of unknown aetiology. We report on three unrelated patients with overlapping de novo interstitial microdeletions involving 5q14.3-q15. All three patients presented with severe psychomotor retardation, epilepsy or febrile seizures, muscular hypotonia and variable brain and minor anomalies. Molecular karyotyping revealed three overlapping microdeletions measuring 5.7, 3.9 and 3.6 Mb, respectively. The microdeletions were identified using single nucleotide polymorphism (SNP) arrays (Affymetrix 100K and Illumina 550K) and array comparative genomic hybridization (1 Mb Sanger array-CGH). Confirmation and segregation studies were performed using fluorescence in situ hybridization (FISH) and quantitative PCR. All three aberrations were confirmed and proven to have occurred de novo. The boundaries and sizes of the deletions in the three patients were different, but an overlapping region of around 1.6 Mb in 5q14.3 was defined. It included five genes: CETN3, AC093510.2, POLR3G, LYSMD3 and the proximal part of GPR98/MASS1, a known epilepsy gene. Haploinsufficiency of GPR98/MASS1 is probably responsible for the seizure phenotype in our patients. At least one other gene contained in the commonly deleted region, LYSMD3, shows a high level of central nervous expression during embryogenesis and is also, therefore, a good candidate gene for other central nervous system (CNS) symptoms, such as psychomotor retardation, brain anomalies and muscular hypotonia of the 5q14.3 microdeletion syndrome.

Stem cell lines that provide a renewable and scaleable supply of central nervous system cell types would constitute an invaluable resource for basic and applied neurobiology. Here we describe the generation and long-term expansion of multiple human foetal neural stem (NS) cell lines in monolayer culture without genetic immortalization. Adherent human NS cells are propagated in the presence of epidermal growth factor (EGF) and fibroblast growth factor 2 (FGF2), under which conditions they stably express neural precursor markers and exhibit negligible differentiation into neurons or glia. However, they produce astrocytes, oligodendrocytes, and neurons upon exposure to appropriate differentiation factors. Single cell cloning demonstrates that human NS cells are tripotent. They retain a diploid karyotype and constant neurogenic capacity after over 100 generations. In contrast to human neurospheres, we observe no requirement for the cytokine leukaemia inhibitory factor (LIF) for continued expansion of adherent human NS cells. Human NS cells can be stably transfected to provide reporter lines and readily imaged in live monolayer cultures, creating the potential for high content genetic and chemical screens.

Background: X-linked ichthyosis (XLI) (steroid sulfatase deficiency) is caused by deletions or point mutations of the steroid sulfatase (STS) gene on chromosome Xp22.32. Deletions of this region can be associated with cognitive behavioural difficulties including autism. Animal work suggests the STS gene may be involved in attentional processes. We have therefore undertaken a systematic study of autism and attention deficit hyperactivity disorder (ADHD) in boys with XLI. Methods: Cases of XLI were recruited from families originally ascertained when pregnancies with STS deficiency were identified through a routine maternal screening programme. Boys with XLI were assessed for ADHD and autism using standardised questionnaires and interviews. Deletions of the STS gene were identified and characterised by analysis of genomic DNA and/or fluorescent in situ hybridisation. Results: 25 boys with XLI were assessed for autism and ADHD. 40% fulfilled DSM-IV criteria for a diagnosis of ADHD, 80% of which were inattentive subtype. ADHD diagnoses were present in those with both deletions and presumed point mutations of STS. Additionally, five boys, from three unrelated families, fulfilled criteria for an autistic spectrum disorder or related language/communication difficulty, and all had an unusually large deletion of the STS gene with loss of the neuroligin 4 (NLGN4) gene. None of the boys with the typical deletion or presumed point mutations of STS demonstrated autistic difficulties. Conclusions: STS deficiency may be a risk factor for ADHD with predominantly inattentive symptoms. Boys with XLI and large deletions encompassing STS and NLGN4 are at increased risk of developing autism and related disorders.

We identified de novo submicroscopic chromosome 14q11.2 deletions in two children with idiopathic developmental delay and cognitive impairment. Vancouver patient 5566 has a approximately 200 kb deletion and Vancouver patient 8326 has a approximately 1.6 Mb deletion. The Database of Chromosomal Imbalance and Phenotype in Humans using Ensembl Resources (DECIPHER) revealed a third patient with idiopathic developmental delay and cognitive impairment, DECIPHER patient 126, who has a approximately 1.1 Mb deletion of 14q11.2. The deletion of patient 5566 overlaps that of patient 126 and both of these deletions lie entirely within that of patient 8326. All three children have similar dysmorphic features, including widely-spaced eyes, short nose with flat nasal bridge, long philtrum, prominent Cupid's bow of the upper lip, full lower lip and similar auricular anomalies.The minimal common deletion region on chromosome 14q11.2 is only approximately 35 kb (from 20.897 to 20.932, University of California at Santa Cruz (UCSC) Genome Browser; build hg18, March 2006) and includes only two genes, SUPT16H and CHD8, which are good candidate genes for the phenotypes. The non-recurrent breakpoints of these patients, the presence of normal copy number variants in the region and the local genomic structure support the notion that this region has reduced stability.

Recently, the application of array-based comparative genomic hybridization (array CGH) has improved rates of detection of chromosomal imbalances in individuals with mental retardation and dysmorphic features. Here, we describe three individuals with learning disability and a heterozygous deletion at chromosome 17q21.3, detected in each case by array CGH. FISH analysis demonstrated that the deletions occurred as de novo events in each individual and were between 500 kb and 650 kb in size. A recently described 900-kb inversion that suppresses recombination between ancestral H1 and H2 haplotypes encompasses the deletion. We show that, in each trio, the parent of origin of the deleted chromosome 17 carries at least one H2 chromosome. This region of 17q21.3 shows complex genomic architecture with well-described low-copy repeats (LCRs). The orientation of LCRs flanking the deleted segment in inversion heterozygotes is likely to facilitate the generation of this microdeletion by means of non-allelic homologous recombination.

Item does not contain fulltext We report the identification of six patients with 3q29 microdeletion syndrome. The clinical phenotype is variable despite an almost identical deletion size. The phenotype includes mild-to-moderate mental retardation, with only slightly dysmorphic facial features that are similar in most patients: a long and narrow face, short philtrum, and high nasal bridge. Autism, gait ataxia, chest-wall deformity, and long and tapering fingers were noted in at least two of six patients. Additional features--including microcephaly, cleft lip and palate, horseshoe kidney and hypospadias, ligamentous laxity, recurrent middle ear infections, and abnormal pigmentation--were observed, but each feature was only found once, in a single patient. The microdeletion is approximately 1.5 Mb in length, with molecular boundaries mapping within the same or adjacent bacterial artificial chromosome (BAC) clones at either end of the deletion in all patients. The deletion encompasses 22 genes, including PAK2 and DLG1, which are autosomal homologues of two known X-linked mental retardation genes, PAK3 and DLG3. The presence of two nearly identical low-copy repeat sequences in BAC clones on each side of the deletion breakpoint suggests that nonallelic homologous recombination is the likely mechanism of disease causation in this syndrome.

We report a case of Feingold syndrome with oesophageal atresia and microcephaly. Marked short stature was present at the age of 9 years. Although short stature has not previously been commented upon as a feature of this syndrome, a review of the literature indicates that it has occurred in several previously reported cases. Of the 18 cases in the literature for which height was recorded, three (16.7%) had height on or below the 0.4th centile, while nine (50%) had height on or below the 10th centile. We suggest that short stature is likely to be a phenotypic feature of Feingold syndrome.

22q11.2 microduplications of a 3-Mb region surrounded by low-copy repeats should be, theoretically, as frequent as the deletions of this region; however, few microduplications have been reported. We show that the phenotype of these patients with microduplications is extremely diverse, ranging from normal to behavioral abnormalities to multiple defects, only some of which are reminiscent of the 22q11.2 deletion syndrome. This diversity will make ascertainment difficult and will necessitate a rapid-screening method. We demonstrate the utility of four different screening methods. Although all the screening techniques give unique information, the efficiency of real-time polymerase chain reaction allowed the discovery of two 22q11.2 microduplications in a series of 275 females who tested negative for fragile X syndrome, thus widening the phenotypic diversity. Ascertainment of the fragile X–negative cohort was twice that of the cohort screened for the 22q11.2 deletion. We also report the first patient with a 22q11.2 triplication and show that this patient's mother carries a 22q11.2 microduplication. We strongly recommend that other family members of patients with 22q11.2 microduplications also be tested, since we found several phenotypically normal parents who were carriers of the chromosomal abnormality.

The type 1 IGF receptor (IGF1R) is required for normal embryonic and postnatal growth. The aim of this study was to determine whether we could detect abnormal IGF1R function in skin fibroblasts from children with an abnormal copy number of the IGF1R gene. We report two children with altered copy number of the IGF1R gene who presented with abnormal growth. Case 1 is a girl with intrauterine growth retardation, postnatal growth failure, and recurrent hypoglycemia. Pituitary function tests were normal. Routine karyotype analysis identified a deletion on 15q26.2, and a fluorescence in situ hybridization study using IGF1R probes showed only a single IGF1R gene. Case 2 was large for gestational age, with birth weight and length at or above 97th percentile, and showed rapid early postnatal growth. He was found to have a recombinant chromosome 15 containing a partial duplication at 15q (q25-qter). A fluorescence in situ hybridization study using the same probes showed three copies of the IGF1R gene. In a mitochondrial activity assay, skin fibroblasts from the subject with only one copy of IGF1R showed slower growth, whereas cells from the subject with three copies of IGF1R showed accelerated growth compared with controls. IGF1R phosphorylation, as assessed by Western blot, and IGF1R binding studies were decreased compared with controls in the child with one copy of the IGF1R and increased in the child with three copies of the gene. Our data are consistent with the concept that IGF1R gene copy number is of functional and clinical importance in humans.

Editor—Five of the human autosomes are acrocentric, chromosomes 13, 14, 15, 21, and 22, and are identified by the presence of satellited short arms. These short arms contain three bands, p11, p12, and p13,1 and are composed of repetitive DNA containing satellite repeats and copies of ribosomal RNA genes. Band p11, the pericentromeric region, is composed of several types of tandemly repeated DNA including satellites I, II, III, and IV, and β satellite DNA.2-4 Band p12, the satellite stalks, contains multiple copies of genes coding for ribosomal RNA5 and is known as the nucleolar organiser region (NOR) as the nucleolus is formed by an aggregation of ribosomal RNA. This can be recognised by staining with silver nitrate (AgNOR staining).6 Band p13 contains β satellite DNA and terminal telomeric sequences.2 7 Loss or gain of the short arm of acrocentric chromosomes occurs without apparent phenotypic effect. For example, Robertsonian translocations occur when two acrocentric chromosomes are joined by centric fusion with the resulting loss of the short arm material and have no associated phenotype in this euchromatically balanced form.8 Chromosomal rearrangements involving the short arms of acrocentric chromosomes are a well known form of chromosomal variation. The most common variation results from rearrangements between the short arms of acrocentric chromosomes. Thus, the satellites of acrocentric chromosomes range in size from no satellites to double or treble satellites as shown by AgNOR staining.8Translocations between the short arm of an acrocentric chromosome and the heterochromatic region of the long arm of the Y chromosome, resulting in acrocentric chromosomes with Y chromosome heterochromatin in place of satellites and satellited Y chromosomes, are also observed.9 10 More rarely, non-acrocentric chromosomes with terminal satellites have been described which arise from a translocation between the short …

A female patient of normal intelligence with short stature and Madelung deformity is reported with Leri-Weill dyschondrosteosis and a de novo pseudodicentric X;Y translocation chromosome. The phenotype is consistent with the observed deletion of the SHOX gene by FISH and molecular studies. The Y chromosome breakpoint was in the short arm but proximal to SRY, consistent with her phenotypic sex. X-inactivation studies have shown a skewed pattern in favour of the dic (X;Y) chromosome. The ARSE gene was also deleted on the dic (X;Y) chromosome but chondrodysplasia punctata was not expressed, as CDP is recessive and ARSE escapes inactivation on the normal X chromosome. Breakpoint mapping assisted in karyotype/phenotype correlation and reproductive counselling. In particular, molecular analysis showed that the putative MRX 49 gene for mental retardation is unlikely to be deleted in this case.

We present seven families with a cytogenetic duplication of the short arm of chromosome 8 at band 8p23.1. The duplication has been transmitted from parents to offspring in four of the seven families. In three families, the source of the extra material and its euchromatic origin were established using FISH with a YAC which was mapped to 8p23.1 and a whole chromosome paint for chromosome 8. FISH signals from this YAC were significantly larger on the duplicated chromosome compared with the normal chromosome in all six family members tested. Comparative genomic hybridisation (CGH) on a representative subject was consistent with these results. The families were ascertained for a variety of mostly incidental reasons including prenatal diagnosis for advanced maternal age. The transmission of this duplication by multiple phenotypically normal family members with no history of reproductive loss suggests the existence of a novel class of 8p23.1 duplications, which can be regarded as euchromatic variants or duplications with no phenotypic effect.

Background Chromosome 15q24 microdeletion syndrome is a rare genomic disorder characterised by intellectual disability, growth retardation, unusual facial morphology and other anomalies. To date, 20 patients have been reported; 18 have had detailed breakpoint analysis. Aim To further delineate the features of the 15q24 microdeletion syndrome, the clinical and molecular characterisation of fifteen patients with deletions in the 15q24 region was performed, nearly doubling the number of reported patients. Methods Breakpoints were characterised using a custom, high-density array comparative hybridisation platform, and detailed phenotype information was collected for each patient. Results Nine distinct deletions with different breakpoints ranging in size from 266 kb to 3.75 Mb were identified. The majority of breakpoints lie within segmental duplication (SD) blocks. Low sequence identity and large intervals of unique sequence between SD blocks likely contribute to the rarity of 15q24 deletions, which occur 8–10 times less frequently than 1q21 or 15q13 microdeletions in our series. Two small, atypical deletions were identified within the region that help delineate the critical region for the core phenotype in the 15q24 microdeletion syndrome. Conclusion The molecular characterisation of these patients suggests that the core cognitive features of the 15q24 microdeletion syndrome, including developmental delays and severe speech problems, are largely due to deletion of genes in a 1.1–Mb critical region. However, genes just distal to the critical region also play an important role in cognition and in the development of characteristic facial features associated with 15q24 deletions. Clearly, deletions in the 15q24 region are variable in size and extent. Knowledge of the breakpoints and size of deletion combined with the natural history and medical problems of our patients provide insights that will inform management guidelines. Based on common phenotypic features, all patients with 15q24 microdeletions should receive a thorough neurodevelopmental evaluation, physical, occupational and speech therapies, and regular audiologic and ophthalmologic screening.

Only a small number of individuals with duplications within the proximal short arm of the X chromosome have been reported. The majority of patients have duplications encompassing Xp11-p21, or extend more distally into Xp22. We report on a female patient who presented within the first year of life with plagiocephaly, speech delay, and epilepsy. Brain MRI showed a relatively thin cerebral cortex, abnormal periventricular white matter, and abnormal vessels in the left inferior parietal region. Cytogenetic and microsatellite analysis of the patient and her parents showed that she has a de novo duplication of Xp11.22-Xp11.4 on her paternal X chromosome. FISH analysis using fluorescently labeled BACs followed by array analysis including an X tilepath BAC array showed that a 12.3 Mb interval between 40.4 Mb and 52.7 Mb from the Xp telomere (NCBI build 36) was duplicated and excluded the presence of additional rearrangements along the X chromosome. Interestingly, X-inactivation studies in peripheral blood leukocytes showed that the duplicated (paternal) X chromosome was active in the majority of cells, in contrast to other patients with Xp duplications in whom X inactivation is random or skewed toward the normal X. These findings suggest that overexpression of genes from proximal Xp is likely to have contributed to her clinical phenotype.

Monoamine oxidases (MAO-A and MAO-B) have a key role in the degradation of amine neurotransmitters, such as dopamine, norepinephrine and serotonin. We identified an inherited 240 kb deletion on Xp11.3-p11.4, which encompasses both monoamine oxidase genes but, unlike other published reports, does not affect the adjacent Norrie disease gene (NDP). The brothers who inherited the deletion, and thus have no monoamine oxidase function, presented with severe developmental delay, intermittent hypotonia and stereotypical hand movements. The clinical features accord with published reports of larger microdeletions and selective MAO-A and MAO-B deficiencies in humans and mouse models and suggest considerable functional compensation between MAO-A and MAO-B under normal conditions.

The 3q29 microdeletion syndrome is caused by a recurrent 1.6 Mb deletion of the 3q subtelomeric region. Though sometimes visible on routine microscopy, the deletion is detected more reliably using subtelomeric fluorescence in-situ hybridization (FISH) or molecular karyotyping. The clinical features associated with a 3q29 microdeletion are variable and include developmental delay, autistic features, skeletal abnormalities and dysmorphic facial features with a relatively long face, long nose with a high bridge and broad tip, short philtrum and large ears. Orofacial clefting, cardiac defects, ocular anomalies and genitourinary malformations have been reported occasionally. We report a three generation family where four individuals were confirmed to have a 3q29 microdeletion and compare their clinical features to those of previously reported patients. This family shows that the learning difficulties associated with a 3q29 deletion may be relatively mild. The history of a severe depressive disorder commencing in adulthood in the affected grandmother also supports previous studies linking the 3q29 region to bipolar disorder and links with the observation of Digilio et al. (2009) who also reported a history of depression in an adult woman with a similar deletion.

Background: Deletions of chromosome 19 have rarely been reported, with the exception of some patients with deletion 19q13.2 and Blackfan–Diamond syndrome due to haploinsufficiency of the RPS19 gene. Such a paucity of patients might be due to the difficulty in detecting a small rearrangement on this chromosome that lacks a distinct banding pattern. Array comparative genomic hybridisation (CGH) has become a powerful tool for the detection of microdeletions and microduplications at high resolution in patients with syndromic mental retardation. Methods and results: Using array CGH, this study identified three interstitial overlapping 19q13.11 deletions, defining a minimal critical region of 2.87 Mb, associated with a clinically recognisable syndrome. The three patients share several major features including: pre- and postnatal growth retardation with slender habitus, severe postnatal feeding difficulties, microcephaly, hypospadias, signs of ectodermal dysplasia, and cutis aplasia over the posterior occiput. Interestingly, these clinical features have also been described in a previously reported patient with a 19q12q13.1 deletion. No recurrent breakpoints were identified in our patients, suggesting that no-allelic homologous recombination mechanism is not involved in these rearrangements. Conclusions: Based on these results, the authors suggest that this chromosomal abnormality may represent a novel clinically recognisable microdeletion syndrome caused by haploinsufficiency of dosage sensitive genes in the 19q13.11 region.

Contains fulltext : 80657.pdf (Publisher’s version ) (Closed access) BACKGROUND: Recurrent 15q13.3 microdeletions were recently identified with identical proximal (BP4) and distal (BP5) breakpoints and associated with mild to moderate mental retardation and epilepsy. METHODS: To assess further the clinical implications of this novel 15q13.3 microdeletion syndrome, 18 new probands with a deletion were molecularly and clinically characterised. In addition, we evaluated the characteristics of a family with a more proximal deletion between BP3 and BP4. Finally, four patients with a duplication in the BP3-BP4-BP5 region were included in this study to ascertain the clinical significance of duplications in this region. RESULTS: The 15q13.3 microdeletion in our series was associated with a highly variable intra- and inter-familial phenotype. At least 11 of the 18 deletions identified were inherited. Moreover, 7 of 10 siblings from four different families also had this deletion: one had a mild developmental delay, four had only learning problems during childhood, but functioned well in daily life as adults, whereas the other two had no learning problems at all. In contrast to previous findings, seizures were not a common feature in our series (only 2 of 17 living probands). Three patients with deletions had cardiac defects and deletion of the KLF13 gene, located in the critical region, may contribute to these abnormalities. The limited data from the single family with the more proximal BP3-BP4 deletion suggest this deletion may have little clinical significance. Patients with duplications of the BP3-BP4-BP5 region did not share a recognisable phenotype, but psychiatric disease was noted in 2 of 4 patients. CONCLUSIONS: Overall, our findings broaden the phenotypic spectrum associated with 15q13.3 deletions and suggest that, in some individuals, deletion of 15q13.3 is not sufficient to cause disease. The existence of microdeletion syndromes, associated with an unpredictable and variable phenotypic outcome, will pose the clinician with diagnostic difficulties and challenge the commonly used paradigm in the diagnostic setting that aberrations inherited from a phenotypically normal parent are usually without clinical consequences.

Background: Genome-wide high resolution array analysis is becoming established as a diagnostic test in the investigation of individuals with learning disability and congenital anomalies; many novel microdeletion and microduplication syndromes have already been identified. The diagnostic use of high resolution array genomic hybridisation analysis for prenatal testing remains to be systematically assessed. Methods: We studied 106 prenatal samples with abnormal ultrasound and a normal karyotype using the Affymetrix GeneChip 6.0 array. “Rare” DNA copy number variations (CNVs) were classified into three groups depending on their size, genomic location and the presence or absence of matched copy number changes in a large cohort of 3000 control samples analysed for copy number changes using genotyping arrays. Results: A total of 35 rare CNVs were identified. 10 (9%) of these are considered likely to represent pathogenic CNVs; 5 were syndromic and 5 were novel. 12 CNVs were detected in at least one control hybridisation and likely to be benign, and 13 CNVs were of unknown clinical significance. In addition, we identified one case of cryptic mosaicism for trisomy 10, one case of loss of heterozygosity (LOH), and showed that the Affymetrix GeneChip 6.0 array platform can detect triploidy. Conclusions: We conclude that careful implementation of high resolution array testing would benefit at least 10% of obstetric patients with abnormal ultrasound findings and a normal karyotype result.

Contains fulltext : 69531.pdf (Publisher’s version ) (Closed access) BACKGROUND: The chromosome 17q21.31 microdeletion syndrome is a novel genomic disorder that has originally been identified using high resolution genome analyses in patients with unexplained mental retardation. AIM: We report the molecular and/or clinical characterisation of 22 individuals with the 17q21.31 microdeletion syndrome. RESULTS: We estimate the prevalence of the syndrome to be 1 in 16,000 and show that it is highly underdiagnosed. Extensive clinical examination reveals that developmental delay, hypotonia, facial dysmorphisms including a long face, a tubular or pear-shaped nose and a bulbous nasal tip, and a friendly/amiable behaviour are the most characteristic features. Other clinically important features include epilepsy, heart defects and kidney/urologic anomalies. Using high resolution oligonucleotide arrays we narrow the 17q21.31 critical region to a 424 kb genomic segment (chr17: 41046729-41470954, hg17) encompassing at least six genes, among which is the gene encoding microtubule associated protein tau (MAPT). Mutation screening of MAPT in 122 individuals with a phenotype suggestive of 17q21.31 deletion carriers, but who do not carry the recurrent deletion, failed to identify any disease associated variants. In five deletion carriers we identify a

PUBLISHED, Background Duplications and deletions in the human genome can cause disease or predispose persons to disease. Advances in technologies to detect these changes allow for the routine identification of submicroscopic imbalances in large numbers of patients. Methods We tested for the presence of microdeletions and microduplications at a specific region of chromosome 1q21.1 in two groups of patients with unexplained mental retardation, autism, or congenital anomalies and in unaffected persons. Results We identified 25 persons with a recurrent 1.35-Mb deletion within 1q21.1 from screening 5218 patients. The microdeletions had arisen de novo in eight patients, were inherited from a mildly affected parent in three patients, were inherited from an apparently unaffected parent in six patients, and were of unknown inheritance in eight patients. The deletion was absent in a series of 4737 control persons (P=1.1x10?7). We found considerable variability in the level of phenotypic expression of the microdeletion; phenotypes included mild-to-moderate mental retardation, microcephaly, cardiac abnormalities, and cataracts. The reciprocal duplication was enriched in nine children with mental retardation or autism spectrum disorder and other variable features (P=0.02). We identified three deletions and three duplications of the 1q21.1 region in an independent sample of 788 patients with mental retardation and congenital anomalies. Conclusions We have identified recurrent molecular lesions that elude syndromic classification and whose disease manifestations must be considered in a broader context of development as opposed to being assigned to a specific disease. Clinical diagnosis in patients with these lesions may be most readily achieved on the basis of genotype rather than phenotype., Supported in part by grants from the National Institutes of Health (NIH) (HD043569, to Dr. Eichler), the South Carolina Department of Disabilities and Special Needs (to Drs. Skinner, Stevenson, and Schwartz), the Wellcome Trust (061183, to Dr. Tassabehji), the Andre & Cyprien Foundation and the University Hospitals of Geneva (to Drs. Antonarakis, Bena, and Gallati), and the European Union (EU) (project 219250, to Dr. Sharp; AnEUploidy project 037627, to Drs. Leeuw, Armengol, Antonarakis, Estivill, Veltman, and de Vries). The Irish Autism Study was funded by the Wellcome Trust and the Health Research Board (a grant to Drs. Gallagher and Gill). Dr. Poot was supported by a grant from the Dutch Foundation for Brain Research (Hersenstichting grant 2008(1) 34); Drs. Regan and Knight, by the Oxford Partnership Comprehensive Biomedical Research Centre; Dr. Willatt, by the Cambridge Biomedical Research Centre, with funding from the United Kingdom Department of Health's National Institute for Health Research Biomedical Research Centres funding scheme; Drs. Huang and Maloney, as part of the National Genetics Reference Laboratory (Wessex) by the United Kingdom Department of Health; Ms. Buysse, as a research assistant of the Research Foundation?Flanders (FWO?Vlaanderen); and Dr. Eichler, as an investigator of the Howard Hughes Medical Institute.

Large-scale copy number variation that is cytogenetically visible in normal individuals has been described as euchromatic variation but needs to be distinguished from pathogenic euchromatic deletion or duplication. Here, we report eight patients (three families and two individuals) with interstitial deletions of 9q13-q21.12. Fluorescence in situ hybridisation with a large panel of BACs showed that all the deleted clones were from extensive tracts of segmentally duplicated euchromatin, copies of which map to both the long and short arms of chromosome 9. The variety of reasons for which these patients were ascertained, and the phenotypically normal parents, indicates that this is a novel euchromatic variant with no phenotypic effect. Further, four patients with classical euchromatic variants of 9q12/qh or 9p12 were also shown to have duplications or triplications of this segmentally duplicated material common to both 9p and 9q. The cytogenetic boundaries between the segmentally duplicated regions and flanking unique sequences were mapped to 9p13.1 in the short arm (BAC RP11-402N8 at 38.7 Mb) and to 9q21.12 in the long arm (BAC RP11-88I18 at 70.3 Mb). The BACs identified in this study should in future make it possible to differentiate between clinically significant deletions or duplications and euchromatic variants with no established phenotypic consequences.

The underlying causes of learning disability and dysmorphic features in many patients remain unidentified despite extensive investigation. Routine karyotype analysis is not sensitive enough to detect subtle chromosome rearrangements (less than 5 Mb). The presence of subtle DNA copy number changes was investigated by array-CGH in 50 patients with learning disability and dysmorphism, employing a DNA microarray constructed from large insert clones spaced at approximately 1 Mb intervals across the genome. Twelve copy number abnormalities were identified in 12 patients (24% of the total): seven deletions (six apparently de novo and one inherited from a phenotypically normal parent) and five duplications (one de novo and four inherited from phenotypically normal parents). Altered segments ranged in size from those involving a single clone to regions as large as 14 Mb. No recurrent deletion or duplication was identified within this cohort of patients. On the basis of these results, we anticipate that array-CGH will become a routine method of genome-wide screening for imbalanced rearrangements in children with learning disability.

We report a de novo, apparently balanced (2;8)(q35;q21.2) translocation in a boy with developmental delay and autism. Cross species (colour) paint (Rx) and SKY FISH, forward and reverse chromosome painting, and FISH with subtelomeric probes were used to examine the patient’s karyotype, but further rearrangements were not detected. FISH with region specific clones mapping near 2q35 and 8q21.2 breakpoints and STS mapping performed on the isolated derivative chromosomes were used to refine the location of the breakpoints further. A cryptic deletion of between 4.23 and 4.41 Mb in extent and involving at least 13 complete genes or transcription units was found at the breakpoint on 2q35. The deletion includes the promoter and 5′ untranslated region of the paired box 3 (PAX3) gene. The child has very mild dystopia canthorum which may be associated with the PAX3 haploinsufficiency. The 8q21.2 breakpoint is within MMP16 which encodes matrix metalloproteinase 16. We postulate that the cryptic deletion and rearrangement are responsible for the patient’s phenotype and that a gene (or genes) responsible for autism lies at 2q35 or 8q21.2. The results present a step towards identifying genes predisposing to autism.

Chromosome 15 is frequently involved in the formation of structural rearrangements. We report the molecular characterisation of 16 independent interstitial duplications, including those of one individual who carried a duplication on both of her chromosomes 15, and three interstitial triplications of the Prader-Willi/Angelman syndrome critical region (PWACR). In all probands except one, the rearrangement was maternal in origin. In one family, the duplication was paternal in origin, yet appeared to segregate in a sibship of three with an abnormal phenotype that included developmental delay and a behavioural disorder. Ten duplications were familial, five de novo and one unknown. All 16 duplications, including two not visible by routine G-banding, were of an almost uniform size and shared the common deletion breakpoints of Prader-Willi syndrome and Angelman syndrome. Like deletions, the formation of duplications can occur in both male and female meiosis and involve both inter- and intrachromosomal events. This implies that at least some deletions and duplications are the reciprocal products of each other. We observed no instances of meiotic instability in the transmission of a duplication, although recombination within the PWACR occurred in two members of the same family between the normal and the duplicated chromosome 15 homologues. All three triplications arose de novo and included alleles from both maternal chromosomes 15. Triplication breakpoints were more variable and extended distally beyond the PWACR. The molecular characteristics of duplications and triplications suggest that they are formed by different mechanisms.

A child with a germline duplication of chromosome 2p, 46,XY,der(13)t(2;13)(p23;q34), who developed a fatal neuroblastoma confirmed at necropsy is reported. Fluorescent in situ hybridisation studies showed chromosome 2p (p23-pter) duplicated on chromosome 13 (q34). The clinical features of the present case shared many similarities to previous reports of trisomy 2p and there have been two cases described with neuroblastoma. Germline duplication of chromosome 2p including the N-myc proto-oncogene may have pre-disposed to the development of neuroblastoma in this case.

This book is a very up to date manual covering the background, methodologies, and applications of molecular cytogenetics techniques. The emphasis is on the diagnostic applications of FISH in the many areas of medicine on which it impinges, including paediatrics, fetal and reproductive medicine, pathology, haematology, oncology, and, of course, medical genetics. With 27 chapters and over 60 authors, all of whom are experts in their field, this book clearly shows how far molecular cytogenetics techniques have developed over the last two decades. The book is divided into three parts. Part 1 covers the basic concepts and techniques. The opening chapter by the editor Yao-Shan Fan provides a very helpful overview of the scope of the book and an extensive set of references for further reading. The following chapters cover probe labelling (DNA and RNA probes) and basic FISH techniques. The second part of the book is devoted to evolving techniques and applications and includes chapters on microdissection, PRINS, SKY FISH, M FISH, CGH, colour banding FISH, fibre FISH, multitelomere FISH, fluorescence genotyping for telomeric regions, and microarray CGH. Special applications …

Editor—Ring chromosomes 2 are rare. We are aware of nine previously reported cases.1-7 We report the clinical and cytogenetic aspects of a further case of ring 2 in a newborn female with severe intrauterine growth retardation (IUGR), microcephaly, heart malformation, and minor dysmorphic features. The subject was born to a 22 year old mother and 30 year old father with no relevant medical history. The mother had two previous miscarriages at 10 and 12 weeks. Her antenatal ultrasound scans in this pregnancy were noted to be abnormal (ventriculomegaly, a two vessel cord, and IUGR). At 34 weeks a repeat ultrasound scan showed reduced liquor volume with no fetal breathing movements. An amniocyte culture was attempted but was unsuccessful. The patient was born by lower segment caesarean section at 35 weeks' gestation. Her birth weight was 1469 g and her head circumference was 29 cm (both below the 3rd centile). She was noted to be dysmorphic and showed severe IUGR. She required ventilation for respiratory distress syndrome. The dysmorphic features include microcephaly, hypertelorism, a wide, flat nasal bridge, bilateral epicanthic folds, narrow and upward slanting palpebral fissures, arched eyebrows, a long philtrum with a thin upper lip, a short neck, right hemihypertrophy, and positional talipes (fig 1). Audiological and ophthalmological assessments have been normal to date. Figure 1 Clinical photograph of the patient. In addition, she has nine cafe au lait patches on her skin, a bifid thumb on her left hand, and a high pitched cry. She had multiple muscular ventriculoseptal defects, which were treated conservatively, and at her most recent review had nearly closed. On follow up until 10 months of age, she has been grossly growth retarded despite an adequate nutrition. Her weight and length are well below the 3rd centile and her head circumference is 4 …

Alveolar capillary dysplasia with misalignment of pulmonary veins (ACD/MPV) is a rare, neonatally lethal developmental disorder of the lung with defining histologic abnormalities typically associated with multiple congenital anomalies (MCA). Using array CGH analysis, we have identified six overlapping microdeletions encompassing the FOX transcription factor gene cluster in chromosome 16q24.1q24.2 in patients with ACD/MPV and MCA. Subsequently, we have identified four different heterozygous mutations (frameshift, nonsense, and no-stop) in the candidate FOXF1 gene in unrelated patients with sporadic ACD/MPV and MCA. Custom-designed, high-resolution microarray analysis of additional ACD/MPV samples revealed one microdeletion harboring FOXF1 and two distinct microdeletions upstream of FOXF1, implicating a position effect. DNA sequence analysis revealed that in six of nine deletions, both breakpoints occurred in the portions of Alu elements showing eight to 43 base pairs of perfect microhomology, suggesting replication error Microhomology-Mediated Break-Induced Replication (MMBIR)/Fork Stalling and Template Switching (FoSTeS) as a mechanism of their formation. In contrast to the association of point mutations in FOXF1 with bowel malrotation, microdeletions of FOXF1 were associated with hypoplastic left heart syndrome and gastrointestinal atresias, probably due to haploinsufficiency for the neighboring FOXC2 and FOXL1 genes. These differences reveal the phenotypic consequences of gene alterations in cis.

Contains fulltext : 34765.pdf (Publisher’s version ) (Closed access) Contains fulltext : 34790.pdf (Publisher’s version ) (Closed access) Array-based comparative genomic hybridization has proven to be successful in the identification of genetic defects in disorders involving mental retardation. Here, we studied a patient with learning disabilities, retinal dystrophy, and short stature. The family history was suggestive of an X-linked contiguous gene syndrome. Hybridization of full-coverage X-chromosomal bacterial artificial chromosome arrays revealed a deletion of ~1 Mb in Xp11.3, which harbors RP2, SLC9A7, CHST7, and two hypothetical zinc-finger genes, ZNF673 and ZNF674. These genes were analyzed in 28 families with nonsyndromic X-linked mental retardation (XLMR) that show linkage to Xp11.3; the analysis revealed a nonsense mutation, p.E118X, in the coding sequence of ZNF674 in one family. This mutation is predicted to result in a truncated protein containing the Kruppel-associated box domains but lacking the zinc-finger domains, which are crucial for DNA binding. We characterized the complete ZNF674 gene structure and subsequently tested an additional 306 patients with XLMR for mutations by direct sequencing. Two amino acid substitutions, p.T343M and p.P412L, were identified that were not found in unaffected individuals. The proline at position 412 is conserved between species and is predicted by molecular modeling to reduce the DNA-binding properties of ZNF674. The p.T343M transition is probably a polymorphism, because the homologous ZNF674 gene in chimpanzee has a methionine at that position. ZNF674 belongs to a cluster of seven highly related zinc-finger genes in Xp11, two of which (ZNF41 and ZNF81) were implicated previously in XLMR. Identification of ZNF674 as the third XLMR gene in this cluster may indicate a common role for these zinc-finger genes that is crucial to human cognitive functioning.