Deep Sequencing and Phenotyping in an Australian Tuberous Sclerosis Complex "No Mutations Identified" Cohort.

Tuberous sclerosis complex (TSC) is a variable multisystem disorder. The "no mutations identified" (NMI) group are reportedly phenotypically milder than those with an identified molecular cause, and often have mosaic or intronic variants not detected by standard sequencing methods. METHODS: We describe the phenotypes in an Australian TSC NMI group (n = 18) and a molecular testing strategy implementable in a diagnostic laboratory. Massively parallel sequencing (MPS) of the whole genomic regions of TSC1 and TSC2 was performed using DNA extracted from multiple tissue samples per participant. RESULTS: Our study showed that the phenotype in TSC NMI individuals can be similar to those with heterozygous, particularly TSC1, variants. Although neurodevelopmental outcomes can be less severe, the number of organ systems involved was similar to the non-mosaic groups. A diagnostic yield of 72% (13/18) was achieved, with the majority (10/13) being mosaic variants and the remainder heterozygous variants missed on previous testing. CONCLUSION: Testing DNA from multiple tissue samples allowed for validation of otherwise discarded low-level mosaic variants and detection of mosaic variants by MPS without excessive cost or the need for specialised techniques. Implementing this approach in a diagnostic setting is viable and allows optimal clinical care of patients with NMI TSC.

Somatic mosaicism in schizophrenia brains reveals prenatal mutational processes.

Germline mutations modulate the risk of developing schizophrenia (SCZ). Much less is known about the role of mosaic somatic mutations in the context of SCZ. Deep (239×) whole-genome sequencing (WGS) of brain neurons from 61 SCZ cases and 25 controls postmortem identified mutations occurring during prenatal neurogenesis. SCZ cases showed increased somatic variants in open chromatin, with increased mosaic CpG transversions (CpG>GpG) and T>G mutations at transcription factor binding sites (TFBSs) overlapping open chromatin, a result not seen in controls. Some of these variants alter gene expression, including SCZ risk genes and genes involved in neurodevelopment. Although these mutational processes can reflect a difference in factors indirectly involved in disease, increased somatic mutations at developmental TFBSs could also potentially contribute to SCZ.

Low-level mosaic trisomy 21 at amniocentesis and cordocentesis in a pregnancy associated with a favorable fetal outcome and perinatal progressive decrease of the trisomy 21 cell line.

OBJECTIVE: We present low-level mosaic trisomy 21 at amniocentesis and cordocentesis in a pregnancy associated with a favorable fetal outcome and perinatal progressive decrease of the trisomy 21 cell line. CASE REPORT: A 36-year-old, primigravid woman underwent amniocentesis at 16 weeks of gestation because of advanced maternal age. This pregnancy was conceived by in vitro fertilization and embryo transfer (IVF-ET). Amniocentesis revealed a karyotype of 47,XY,+21 [3]/46,XY [17] (15% mosaicism) and simultaneous array comparative genomic hybridization (aCGH) analysis on the DNA extracted from uncultured amniocytes revealed the result of arr (21) × 2∼3 (X,Y) × 1, consistent with 24.5% mosaicism for trisomy 21. Cordocentesis performed at 21 weeks of gestation revealed a karyotype of 47,XY,+21 [3]/46,XY [37] (6% mosaicism). She was referred for genetic counseling at 31 weeks of gestation, and continuing the pregnancy was advised. The parental karyotypes and prenatal ultrasound were normal. At 37 weeks of gestation, a phenotypically normal baby was delivered with a body weight of 2900-g. The karyotypes of cord blood, umbilical cord and placenta were 47,XY,+21 [1]/46,XY [39] (2.5% mosaicism), 47,XY,+21 [10]/46,XY [30] (25% mosaicism) and 47,XY,+21 [22]/46,XY [18] (55% mosaicism), respectively. Quantitative fluorescent polymerase chain reaction (QF-PCR) analysis on the DNA extracted from umbilical cord and parental bloods excluded uniparental disomy (UPD) 21 and revealed a maternal origin of the extra chromosome 21. When follow-up at the age of 2 months, the neonate was normal in phenotype and development. The peripheral blood had a karyotype of 47,XY,+21 [1]/46,XY [39] (2.5% mosaicism), and interphase fluorescence in situ hybridization (FISH) analysis on uncultured buccal mucosal cells revealed 4.7% (5/105 cells) mosaicism for trisomy 21, compared with 0% (5/100 cells) in the normal control. CONCLUSION: Low-level mosaic trisomy 21 at amniocentesis and cordocentesis can be associated with favorable fetal outcome and perinatal progressive decrease of the trisomy 21 cell line.

Low-level mosaic trisomy 21 due to mosaic unbalanced Robertsonian translocation of 46,XX,+21,der(21;21) (q10;q10)/46,XX at amniocentesis in a pregnancy associated with a favorable fetal outcome, cytogenetic discrepancy between cultured amniocytes and uncultured amniocytes, cytogenetic discrepancy among various tissues and perinatal progressive decrease of the trisomy 21 cell line.

OBJECTIVE: We present prenatal diagnosis of mosaic trisomy 21 at amniocentesis associated with unbalanced Robertsonian translocation in the fetus and a favorable fetal outcome. CASE REPORT: A 41-year-old, primigravid woman underwent amniocentesis at 17 weeks of gestation because of advanced maternal age. Her husband was 41 years old. Amniocentesis revealed a karyotype of 46,XX,+21,der(21;21) (q10;q10)[8]/46,XX[18], consistent with 30.8% (8/26 colonies) mosaicism for trisomy 21. Simultaneous array comparative genomic hybridization (aCGH) analysis on the DNA extracted from uncultured amniocytes revealed the result of arr (1-22,X) × 2 with no genomic imbalance. Repeat amniocentesis at 21 weeks of gestation revealed a karyotype of 46,XX,+21,der(21;21) (q10;q10)[2]/46,XX[25], consistent with 7.4% (2/27 colonies) mosaicism for trisomy 21. Cord blood sampling revealed the result of 46,XX and rsa X(P095) × 2, 13,18,21(P095) × 2. Prenatal ultrasound findings were normal. At 23 weeks of gestation, she underwent cord blood sampling which revealed a karyotype of 46,XX. At 26 weeks of gestation, she was referred for genetic counseling. No repeat amniocentesis and continuing the pregnancy were advised. The mother had a karyotype of 46,XX, and the father had a karyotype of 46,XY. At 38 weeks of gestation, a 3476-g, phenotypically normal baby was delivered. The cord blood had a karyotype of 46,XX,+21,der(21;21) (q10;q10)[1]/46,XX[39] (2.5% mosaicism). The placenta had a karyotype of 46,XX,+21,der(21;21) (q10;q10) (40/40 cells). When follow-up at age two months, the neonate was normal in phenotype and development. The peripheral blood had a karyotype of 46,XX (40/40 cells), and aCGH analysis on buccal mucosal cells resulted no genomic imbalance. CONCLUSION: Low-level mosaic trisomy 21 at amniocentesis due to mosaic unbalanced Robertsonian translocation with a normal cell line can be associated with a favorable fetal outcome, cytogenetic discrepancy between cultured amniocytes and uncultured amniocytes, cytogenetic discrepancy among various tissues and perinatal progressive decrease of the trisomy 21 cell line.

The Clinical Spectrum of Mosaic Genetic Disease.

Genetic mosaicism is defined as the presence of two or more cell lineages with different genotypes arising from a single zygote. Mosaicism has been implicated in hundreds of genetic diseases with diverse genetic etiologies affecting every organ system. Mosaic genetic disease (MDG) is a spectrum that, on the extreme ends, enables survival from genetic severe disorders that would be lethal in a non-mosaic form. On the milder end of the spectrum, mosaicism can result in little if any phenotypic effects but increases the risk of transmitting a pathogenic genotype. In the middle of the spectrum, mosaicism has been implicated in reducing the phenotypic severity of genetic disease. In this review will describe the spectrum of mosaic genetic disease whilst discussing the status of the detection and prevalence of mosaic genetic disease.

Association of autosomal mosaic chromosomal alterations with risk of bladder cancer in Chinese adults: a prospective cohort study.

Little is known about the prospective association between autosomal mosaic chromosomal alterations (mCAs), a group of large-scale somatic mutations on autosomes, and bladder cancer. Here we utilized data from 99,877 participants who were free of physician-diagnosed cancer at baseline (2004-2008) of the China Kadoorie Biobank to estimate the associations between autosomal mCAs and bladder cancer (ICD-10: C67). A total of 2874 autosomal mCAs events among 2612 carriers (2.6%) were detected. After a median follow-up of 12.4 years, we discovered that participants with all autosomal mCAs exhibited higher risks of bladder cancer, with a multivariable-adjusted hazard ratio (HR) (95% confidence interval [CI]) of 2.60 (1.44, 4.70). The estimate of such association was even stronger for mosaic loss events (HR [95% CI]: 6.68 [2.92, 15.30]), while it was not significant for CN-LOH events. Both expanded (cell fraction ≥10%) and non-expanded autosomal mCAs, as well as mosaic loss, were associated with increased risks of bladder cancer. Of interest, physical activity (PA) significantly modified the associations of autosomal mCAs and mosaic loss (Pinteraction = 0.038 and 0.012, respectively) with bladder cancer. The increased risks of bladder cancer were only observed with mCAs and mosaic loss among participants with a lower level of PA (HR [95% CI]: 5.11 [2.36, 11.09] and 16.30 [6.06, 43.81]), but not among participants with a higher level of PA. Our findings suggest that peripheral leukocyte autosomal mCAs may represent a novel risk factor for bladder cancer, and PA may serve as a potential intervention target for mCAs carriers.

Slc35a2 mosaic knockout impacts cortical development, dendritic arborisation, and neuronal firing.

Mild malformation of cortical development with oligodendroglial hyperplasia in epilepsy (MOGHE) is an important cause of drug-resistant epilepsy. A significant subset of individuals diagnosed with MOGHE display somatic mosaicism for loss-of-function variants in SLC35A2, which encodes the UDP-galactose transporter. We developed a mouse model to investigate how disruption of this transporter leads to a malformation of cortical development. We used in utero electroporation and CRISPR/Cas9 to knockout Slc35a2 in a subset of layer 2/3 cortical neuronal progenitors in the developing brains of male and female fetal mice to model mosaic expression. Mosaic Slc35a2 knockout was verified through next-generation sequencing and immunohistochemistry of GFP-labelled transfected cells. Histology of brain tissue in mosaic Slc35a2 knockout mice revealed the presence of upper layer-derived cortical neurons in the white matter. Reconstruction of single filled neurons identified altered dendritic arborisation with Slc35a2 knockout neurons having increased complexity. Whole-cell electrophysiological recordings revealed that Slc35a2 knockout neurons display reduced action potential firing, increased afterhyperpolarisation duration and reduced burst-firing when compared with control neurons. Mosaic Slc35a2 knockout mice also exhibited significantly increased epileptiform spiking and increased locomotor activity. We successfully generated a mouse model of mosaic Slc35a2 deficiency, which recapitulates features of the human phenotype, including impaired neuronal migration. We show that knockout in layer 2/3 cortical neuron progenitors is sufficient to disrupt neuronal excitability, increase epileptiform activity and cause hyperactivity in mosaic mice. Our mouse model provides an opportunity to further investigate the disease mechanisms that contribute to MOGHE and facilitate the development of precision therapies.

Genetic counseling of mosaicism for a duplication due to partial trisomy in a cell line with 46 chromosomes associated with a normal cell line at amniocentesis.

Genetic counseling of mosaicism for a duplication due to partial trisomy in a cell line with 46 chromosomes associated with a normal cell line at amniocentesis remains difficult because mosaic duplication due to partial trisomy has been reported to be associated with either normal or abnormal phenotype in prenatal diagnosis. This article makes a comprehensive review of the reported cases of mosaicism for a duplication due to partial trisomy in a cell line with 46 chromosomes associated with a normal cell line at amniocentesis and various counseling issues such as culture artefact, cytogenetic discrepancy between cultured and uncultured amniocytes and among various tissues, perinatal progressive decrease of the abnormal cell line and a possible favorable fetal outcome. The information provided is useful for obstetricians and genetic counselors during genetic counseling of the parents who wish to keep the babies under such a circumstance.

Genetic counseling of mosaic and non-mosaic tetrasomy 9p at prenatal diagnosis.

Genetic counseling of mosaic and non-mosaic tetrasomy 9p remains difficult because of the possible associated congenital abnormalities, cytogenetic discrepancy in various tissues, true-positive and false-positive diagnosis in non-invasive prenatal testing (NIPT), uniparental disomy (UPD) 9, tissue-limited mosaicism, perinatal progressive decrease of the aneuploid cell line, phenotypic normal carriers and possible favorable fetal outcome in the cases with mosaic tetrasomy 9p at amniocentesis. This article presents a comprehensive review of various counseling issues concerning mosaic and non-mosaic tetrasomy 9p at prenatal diagnosis, and the information provided is very useful for genetic counseling under such circumstances.

Genetic counseling of mosaicism for balanced or unbalanced translocation with a normal cell line at amniocentesis.

Genetic counseling of mosaicism for balanced translocation with a normal cell line at amniocentesis is not difficult because most of the reported cases have normal phenotypes. However, genetic counseling of mosaicism for unbalanced translocation with a normal cell line at amniocentesis remains difficult because cases with mosaic unbalanced translocation with a normal cell line at prenatal diagnosis have been reported to be associated with either normal or abnormal phenotype. This article makes a comprehensive review of the reported cases of de novo or familial mosaic unbalanced translocation with a normal cell line and various counseling issues such as meiotic event, post-zygotic mitotic event, culture artefact, chimerism, uniparental disomy (UPD), jumping translocation, cytogenetic discrepancy between cultured and uncultured amniocytes and among various tissues, perinatal progressive decrease of the unbalanced translocation cell line and a possible favorable fetal outcome. The information provided is useful for obstetricians and genetic counselors during genetic counseling of the parents who wish to keep the babies under such a circumstance.

Genetic counseling of mosaicism for a deletion due to partial monosomy in a cell line with 46 chromosomes associated with a normal cell line at amniocentesis.

Genetic counseling of mosaicism for a deletion due to partial monosomy in a cell line with 46 chromosomes associated with a normal cell line at amniocentesis remains difficult because mosaic deletion due to partial monosomy has been reported to be associated with either normal or abnormal phenotype in prenatal diagnosis. This article makes a comprehensive review of the reported cases of mosaicism for a deletion due to partial monosomy in a cell line with 46 chromosomes associated with a normal cell line at amniocentesis and various counseling issues such as culture artefact, cytogenetic discrepancy between cultured and uncultured amniocytes and among various tissues, perinatal progressive decrease of the abnormal cell line and a possible favorable fetal outcome. The information provided is useful for obstetricians and genetic counselors during genetic counseling of the parents who wish to keep the babies under such a circumstance.

Low-level mosaic trisomy 14 at amniocentesis in a pregnancy associated with cytogenetic discrepancy between cultured amniocytes and uncultured amniocytes, positive non-invasive prenatal testing for trisomy 14, perinatal progressive decrease of the trisomy 14 cell line and a favorable fetal outcome.

OBJECTIVE: We present low-level mosaic trisomy 14 at amniocentesis. CASE REPORT: A 37-year-old, gravida 2, para 1, woman underwent amniocentesis at 18 weeks of gestation because of advanced maternal age. This pregnancy was conceived by in vitro fertilization and embryo transfer (IVF-ET). Amniocentesis revealed a karyotype of 47,XX,+14 [4]/46,XX [27], consistent with 12.9% mosaicism for trisomy 14. Simultaneous array comparative genomic hybridization (aCGH) analysis on the DNA extracted from uncultured amniocytes revealed the result of arr (1-22, X) × 2 with no genomic imbalance. Prenatal ultrasound findings were unremarkable. She was referred for genetic counseling at 21 weeks of gestation and was offered expanded non-invasive prenatal testing (NIPT) which was positive for trisomy 14. At 24 weeks of gestation, she underwent repeat amniocentesis which revealed a karyotype of 47,XX,+14 [2]/46,XX [26], consistent with 7% mosaicism for trisomy 14. The parental karyotypes were normal. Simultaneous aCGH analysis on the DNA extracted from uncultured amniocytes revealed no genomic imbalance. Polymorphic marker analysis excluded uniparental disomy (UPD) 14. Interphase fluorescence in situ hybridization (FISH) analysis on 104 uncultured amniocytes detected no trisomy 14 cell. At 35 weeks of gestation, a 2315-g phenotypically normal baby was delivered. The umbilical cord and placenta had the karyotype of 46, XX (40/40 cells). aCGH analysis on the DNA extracted from peripheral blood and buccal mucosal cells at the age of three months revealed no genomic imbalance. The neonate was normal in phenotype and development during postnatal follow-ups. CONCLUSIONS: Low-level mosaic trisomy 14 at amniocentesis can be associated with cytogenetic discrepancy between cultured amniocytes and uncultured amniocytes, perinatal progressive decrease of the trisomy 14 cell line and a favorable fetal outcome.

Mosaic distal 13q duplication due to mosaic unbalanced translocation of 46,XY,der(14)t(13;14)(q32.2;p13)/46,XY at amniocentesis in a pregnancy associated with a favorable fetal outcome, perinatal progressive decrease of the aneuploid cell line and cytogenetic discrepancy between cultured amniocytes and uncultured amniocytes.

OBJECTIVE: We present mosaic distal 13q duplication due to mosaic unbalanced translocation 46,XY,der(14)t(13;14)(q32.2;p13)/46,XY at amniocentesis in a pregnancy associated with a favorable fetal outcome. CASE REPORT: A 37-year-old, gravida 2, para 0, woman underwent amniocentesis at 17 weeks of gestation because of advanced maternal age. Amniocentesis revealed a karyotype of 46,XY, add(14) (p13)[17]/46,XY[13] (56.6% mosaicism). Array comparative genomic hybridization (aCGH) analysis on the DNA extracted from cultured amniocytes revealed arr 13q32.2q34 × 2∼3, consistent with 45% mosaicism for distal 13q duplication. Repeat amniocentesis at 24 weeks of gestation revealed a karyotype of 46,XY,der(14)t(13;14)(q32.2;p13)[14]/46,XY[16] (46.6% mosaicism). The parental karyotypes were normal. aCGH analysis on the DNA extracted from uncultured amniocytes revealed arr 13q32.2q34 × 2.38, consistent with 30-40% mosaicism for distal 13q duplication. Interphase fluorescence in situ hybridization (FISH) analysis on uncultured amniocytes detected 22.8% (23/101 cells) mosaicism for distal 13q duplication. Prenatal ultrasound findings were unremarkable. At 39 weeks of gestation, a 3616-g phenotypically normal baby was delivered. The karyotypes of cord blood, umbilical cord and placenta were 46,XY,der(14)t(13;14)(q32.2;p13)[20]/46,XY[20] (50% mosaicism), 46,XY,der(14)t(13;14)(q32.2;p13)[14]/46,XY[26] (35% mosaicism) and 46,XY (40/40 cells) (0% mosaicism), respectively. When follow-ups at the age of 4½ months and the age of one year, the peripheral blood had the karyotype of 46,XY,der(14)t(13;14)(q32.2;p13)[18]/46,XY[22] (45% mosaicism). Interphase FISH analysis on buccal mucosal cells at the age of 4½ months revealed 2.7% (3/110 cells) mosaicism for distal 13q duplication, compared with 1% (1/100 cells) in the normal control. The neonate was normal in phenotype and development. CONCLUSIONS: Mosaic unbalanced translocation at amniocentesis can be associated with a favorable fetal outcome, perinatal progressive decrease of the aneuploid cell line and cytogenetic discrepancy between cultured amniocytes and uncultured amniocytes.

Prenatal diagnosis and molecular cytogenetic analyses of a homozygous Robertsonian translocation family with novel mosaic Robertsonian fission karyotype.

BACKGROUND: Approximately one person in 1000 is a Robertsonian translocation carrier. Errors in the formation of eggs (or more rarely of sperms) may be the cause of Robertsonian translocation. Most Robertsonian translocation carriers are healthy and have a normal lifespan, but do have an increased risk of offsprings with trisomies and pregnancy loss. The fitness of Robertsonian translocation carriers is reduced, but can provide material for evolution. MATERIALS AND METHODS: We have done prenatal diagnosis and molecular cytogenetic analyses on this homozygous Robertson translocation family. We report a homozygous Robertson translocation family with previously undescribed mosaic Robertsonian fission karyotype. RESULTS: We identified six Robertsonian translocation carriers in this family. Four were heterozygous translocation carriers of 45,XX or XY,der(14;15)(q10;q10), one was a homozygous translocation carrier of a 44,XY,der(14;15)(q10;q10),der(14;15)(q10;q10), and one was a previously undescribed Robertsonian fission carrier of 45,XN,der(14;15)(q10;q10)[42]/46,XN[58] with normal phenotype. CONCLUSION: We reported a previously undescribed mosaic Robertsonian fission karyotype. The homozygosity of Robertsonian translocation for speciation may be a potential mechanism of speciation in humans. In theory, the carriers of homologous Robertsonian translocation cannot produce normal gametes, but Robertson fission made it possible for them to produce normal gametes.

Case report: Deep sequencing and long-read genome sequencing refine prior genetic analyses in families with apparent gonadal mosaicism in PIK3CD-related activated PI3K delta syndrome.

Gonadal and gonosomal mosaicism describe phenomena in which a seemingly healthy individual carries a genetic variant in a subset of their gonadal tissue or gonadal and somatic tissue(s), respectively, with risk of transmitting the variant to their offspring. In families with one or more affected offspring, occurrence of the same apparently de novo variants can be an indicator of mosaicism in either parent. Panel-based deep sequencing has the capacity to detect low-level mosaic variants with coverage exceeding the typical limit of detection provided by current, readily available sequencing techniques. In this study, we report three families with more than one affected offspring with either confirmed or apparent parental gonosomal or gonadal mosaicism for PIK3CD pathogenic variants. Data from targeted deep sequencing was suggestive of low-level maternal gonosomal mosaicism in Family 1. Through this approach we did not detect pathogenic variants in PIK3CD from parental samples in Family 2 and Family 3. We conclude that mosaicism was likely confined to the maternal gonads in Family 2. Subsequent long-read genome sequencing in Family 3 showed that the paternal chromosome harbored the pathogenic variant in PIK3CD in both affected children, consistent with paternal gonadal mosaicism. Detection of parental mosaic variants enables accurate risk assessment, informs reproductive decision-making, and provides helpful context to inform clinical management in families with PIK3CD pathogenic variants.

Revertant Mosaic Skin Punch Grafting in Recessive Dystrophic Epidermolysis Bullosa.

This case report describes a patient with severe generalized recessive dystrophic epidermolysis bullosa who presented with an unhealed ulcer that had persisted for the past 3 years.

The impact of mosaic loss of the Y chromosome (mLOY) in men of advanced age.

The Y chromosome has long been considered to be a "genetic wasteland" harboring only few genes essentially involved in male sex development and spermatogenesis. However, the discovery of mosaic loss of the Y chromosome (mLOY) in older men has led to revisiting of the potential impact of the Y chromosome on health and the pathophysiological processes of multiple diseases such as cancer, Alzheimer's disease and cardiovascular disease. Hence, developing more sensitive techniques for the detection of mLOY has become an emergent concern. In this article, we present a comprehensive review of the literature regarding mLOY. Additionally, we discuss the emerging discoveries concerning mLOY as well as the underlying mechanisms promoting disease in men of advanced age.