Growth and neurodevelopmental disorder with arthrogryposis, microcephaly and structural brain anomalies caused by Bi-allelic partial deletion of SMPD4 gene.

Neutral sphingomyelinases have an important role in generation of ceramide and phosphorylcholine from sphingomyelins which then act as secondary messengers in various signaling pathways of the cellular machinery. They function ubiquitously with a predominant role in the central nervous system. Neutral sphingomyelinase type 3, encoded by SMPD4 gene has recently been reported to cause a severe autosomal recessive neurodevelopmental disorder with congenital arthrogryposis and microcephaly. We report a 22-month-old girl having characteristic features of neurodevelopmental delay, prenatal onset growth failure, arthrogryposis, microcephaly and brain anomalies including severe hypomyelination, simplified gyral pattern and hypoplasia of corpus callosum and brain stem. In addition, she was noted to have nystagmus and visual impairment secondary to macular dystrophy and retinal pigment epithelial stippling at posterior pole. Copy number variant analysis from trio whole exome sequencing (ES) enabled identification of a homozygous 11 kb deletion encompassing exons 18-20 of SMPD 4 gene, confirming the diagnosis of SMPD4-related disorder in her.

A data set of variants derived from 1455 clinical and research exomes is efficient in variant prioritization for early-onset monogenic disorders in Indians.

Given the genomic uniqueness, a local data set is most desired for Indians, who are underrepresented in existing public databases. We hypothesize patients with rare monogenic disorders and their family members can provide a reliable source of common variants in the population. Exome sequencing (ES) data from families with rare Mendelian disorders was aggregated from five centers in India. The dataset was refined by excluding related individuals and removing the disease-causing variants (refined cohort). The efficiency of these data sets was assessed in a new set of 50 exomes against gnomAD and GenomeAsia. Our original cohort comprised 1455 individuals from 1203 families. The refined cohort had 836 unrelated individuals that retained 1,251,064 variants with 181,125 population-specific and 489,618 common variants. The allele frequencies from our cohort helped to define 97,609 rare variants in gnomAD and 44,520 rare variants in GenomeAsia as common variants in our population. Our variant dataset provided an additional 1.7% and 0.1% efficiency for prioritizing heterozygous and homozygous variants respectively for rare monogenic disorders. We observed additional 19 genes/human knockouts. We list carrier frequency for 142 recessive disorders. This is a large and useful resource of exonic variants for Indians. Despite limitations, datasets from patients are efficient tools for variant prioritization in a resource-limited setting.

Functional characterization of novel variants in SMPD1 in Indian patients with acid sphingomyelinase deficiency.

Pathogenic variations in SMPD1 lead to acid sphingomyelinase deficiency (ASMD), that is, Niemann-Pick disease (NPD) type A and B (NPA, NPB), which is a recessive lysosomal storage disease. The knowledge of variant spectrum in Indian patients is crucial for early and accurate NPD diagnosis and genetic counseling of families. In this study, we recruited 40 unrelated pediatric patients manifesting symptoms of ASMD and subnormal ASM enzyme activity. Variations in SMPD1 were studied using Sanger sequencing for all exons, followed by interpretation of variants based on American College of Medical Genetics and Genomics & Association for Molecular Pathology (ACMG/AMP) criteria. We identified 18 previously unreported variants and 21 known variants, including missense, nonsense, deletions, duplications, and splice site variations with disease-causing potential. Eight missense variants were functionally characterized using in silico molecular dynamic simulation and in vitro transient transfection in HEK293T cells, followed by ASM enzyme assay, immunoblot, and immunofluorescence studies. All the variants showed reduced ASM activity in transfected cells confirming their disease-causing potential. The study provides data for efficient prenatal diagnosis and genetic counseling of families with NPD type A and B.

Clinical and genetic spectrum of 104 Indian families with central nervous system white matter abnormalities.

Genetic disorders with predominant central nervous system white matter abnormalities (CNS WMAs), also called leukodystrophies, are heterogeneous entities. We ascertained 117 individuals with CNS WMAs from 104 unrelated families. Targeted genetic testing was carried out in 16 families and 13 of them received a diagnosis. Chromosomal microarray (CMA) was performed for three families and one received a diagnosis. Mendeliome sequencing was used for testing 11 families and all received a diagnosis. Whole exome sequencing (WES) was performed in 80 families and was diagnostic in 52 (65%). Singleton WES was diagnostic for 50/75 (66.67%) families. Overall, genetic diagnoses were obtained in 77 families (74.03%). Twenty-two of 47 distinct disorders observed in this cohort have not been reported in Indian individuals previously. Notably, disorders of nuclear mitochondrial pathology were most frequent (9 disorders in 20 families). Thirty-seven of 75 (49.33%) disease-causing variants are novel. To sum up, the present cohort describes the phenotypic and genotypic spectrum of genetic disorders with CNS WMAs in our population. It demonstrates WES, especially singleton WES, as an efficient tool in the diagnosis of these heterogeneous entities. It also highlights possible founder events and recurrent disease-causing variants in our population and their implications on the testing strategy.

NGS-based expanded carrier screening for genetic disorders in North Indian population reveals unexpected results - a pilot study.

BACKGROUND: To determine the carrier frequency and pathogenic variants of common genetic disorders in the north Indian population by using next generation sequencing (NGS). METHODS: After pre-test counselling, 200 unrelated individuals (including 88 couples) were screened for pathogenic variants in 88 genes by NGS technology. The variants were classified as per American College of Medical Genetics criteria. Pathogenic and likely pathogenic variants were subjected to thorough literature-based curation in addition to the regular filters. Variants of unknown significance were not reported. Individuals were counselled explaining the implications of the results, and cascade screening was advised when necessary. RESULTS: Of the 200 participants, 52 (26%) were found to be carrier of one or more disorders. Twelve individuals were identified to be carriers for congenital deafness, giving a carrier frequency of one in 17 for one of the four genes tested (SLC26A4, GJB2, TMPRSS3 and TMC1 in decreasing order). Nine individuals were observed to be carriers for cystic fibrosis, with a frequency of one in 22. Three individuals were detected to be carriers for Pompe disease (frequency one in 67). None of the 88 couples screened were found to be carriers for the same disorder. The pathogenic variants observed in many disorders (such as deafness, cystic fibrosis, Pompe disease, Canavan disease, primary hyperoxaluria, junctional epidermolysis bullosa, galactosemia, medium chain acyl CoA deficiency etc.) were different from those commonly observed in the West. CONCLUSION: A higher carrier frequency for genetic deafness, cystic fibrosis and Pompe disease was unexpected, and contrary to the generally held view about their prevalence in Asian Indians. In spite of the small sample size, this study would suggest that population-based carrier screening panels for India would differ from those in the West, and need to be selected with due care. Testing should comprise the study of all the coding exons with its boundaries in the genes through NGS, as all the variants are not well characterized. Only study of entire coding regions in the genes will detect carriers with adequate efficiency, in order to reduce the burden of genetic disorders in India and other resource poor countries.

Expanding the phenotypic and genotypic spectrum of Wiedemann-Steiner syndrome: First patient from India.

Wiedemann-Steiner syndrome (WWS) is a rare disorder characterized by hypotonia, postnatal growth restriction, striking facial dysmorphism, and hirsutism. It is caused by heterozygous pathogenic variants in KMT2A. This gene has an established role in histone methylation, which explains the overlap of WWS with syndromes caused by genes involved in chromatin remodeling. We describe an infant with a novel single base pair deletion in KMT2A with features consistent with WWS, as well as additional features of stenosis of aqueduct of Sylvius and broad toes. The usefulness of Face2Gene as a tool for identification of dysmorphology syndromes is discussed, as in this patient, it suggested WWS as the top candidate disorder. To the best of our knowledge, this is the first patient of WWS reported from India, with a novel genotype and expanded phenotype.

The first case of antenatal presentation in COG8-congenital disorder of glycosylation with a novel splice site mutation and an extended phenotype.

Congenital disorders of glycosylation (CDG) are an extremely rapidly growing and phenotypically versatile group of disorders. Conserved oligomeric Golgi (COG) complexes are hetero-octameric proteins involved in retrograde trafficking within the Golgi. Seven of its eight subunits have a causal role in CDG. To date, only three cases of COG8-CDG have been published but none in the antenatal period. We present the first case of antenatally diagnosed COG8-CDG with facial dysmorphism and additional features such as Dandy-Walker malformation and arthrogryposis multiplex congenita, thus expanding the phenotype of this rare disorder. Trio whole exome sequencing revealed a novel homozygous variant in COG8, which creates a new splice site in exon 5 and protein truncation after 12 amino acids downstream to the newly generated splice site. As the mutations of the previous three patients were also identified in exon 5, it is likely to be a potential mutational hotspot in COG8. An association between antenatally increased nuchal translucency and COG8-CDG is also established, which would alert clinicians to its diagnosis early in gestation. It remains to be seen if this observation can be extended to other COG-CDGs.

Clinical and molecular characterization of Indian patients with fructose-1, 6-bisphosphatase deficiency: Identification of a frequent variant (E281K).

Fructose-1, 6-bisphosphatase deficiency is an autosomal recessive disorder of gluconeogenesis caused by genetic defect in the FBP1 gene. It is characterized by episodic, often life-threatening metabolic acidosis, liver dysfunction, and hyperlactatemia. Without a high index of suspicion, it may remain undiagnosed with devastating consequences. Accurate diagnosis can be achieved either by enzyme assay or gene studies. Enzyme assay requires a liver biopsy and is tedious, invasive, expensive, and not easily available. Therefore, genetic testing is the most appropriate method to confirm the diagnosis. Molecular studies were performed on 18 suspected cases presenting with episodic symptoms. Seven different pathogenic variants were identified. Two common variants were noted in two subpopulations from the Indian subcontinent; p.Glu281Lys (E281K) occurred most frequently (in 10 patients) followed by p.Arg158Trp (R158W, in 4 patients). Molecular analysis confirmed the diagnosis and helped in managing these patients by providing appropriate genetic counseling. In conclusion, genetic studies identified two common variants in the Indian subcontinent, thus simplifying the diagnostic algorithm in this treatable disorder.

LYRM7 mutations cause a multifocal cavitating leukoencephalopathy with distinct MRI appearance.

This study focused on the molecular characterization of patients with leukoencephalopathy associated with a specific biochemical defect of mitochondrial respiratory chain complex III, and explores the impact of a distinct magnetic resonance imaging pattern of leukoencephalopathy to detect biallelic mutations in LYRM7 in patients with biochemically unclassified leukoencephalopathy. 'Targeted resequencing' of a custom panel including genes coding for mitochondrial proteins was performed in patients with complex III deficiency without a molecular genetic diagnosis. Based on brain magnetic resonance imaging findings in these patients, we selected additional patients from a database of unclassified leukoencephalopathies who were scanned for mutations in LYRM7 by Sanger sequencing. Targeted sequencing revealed homozygous mutations in LYRM7, encoding mitochondrial LYR motif-containing protein 7, in four patients from three unrelated families who had a leukoencephalopathy and complex III deficiency. Two subjects harboured previously unreported variants predicted to be damaging, while two siblings carried an already reported pathogenic homozygous missense change. Sanger sequencing performed in the second cohort of patients revealed LYRM7 mutations in three additional patients, who were selected on the basis of the magnetic resonance imaging pattern. All patients had a consistent magnetic resonance imaging pattern of progressive signal abnormalities with multifocal small cavitations in the periventricular and deep cerebral white matter. Early motor development was delayed in half of the patients. All patients but one presented with subacute neurological deterioration in infancy or childhood, preceded by a febrile infection, and most patients had repeated episodes of subacute encephalopathy with motor regression, irritability and stupor or coma resulting in major handicap or death. LYRM7 protein was strongly reduced in available samples from patients; decreased complex III holocomplex was observed in fibroblasts from a patient carrying a splice site variant; functional studies in yeast confirmed the pathogenicity of two novel mutations. Mutations in LYRM7 were previously found in a single patient with a severe form of infantile onset encephalopathy. We provide new molecular, clinical, and neuroimaging data allowing us to characterize more accurately the molecular spectrum of LYRM7 mutations highlighting that a distinct and recognizable magnetic resonance imaging pattern is related to mutations in this gene. Inter- and intrafamilial variability exists and we observed one patient who was asymptomatic by the age of 6 years.

Is the diagnostic yield influenced by the indication for fetal autopsy?

The utility of fetal autopsy to corroborate antenatal ultrasound findings and to aid genetic counseling is well known. However, the ability to identify an underlying cause for the common indications for which it is performed is not well studied. This study aimed to determine if the diagnostic yield of fetal autopsy in identifying the underlying cause is determined by the indication of the autopsy. Five groups of fetuses were defined based on the indication for the autopsy performed in 903 cases: (i) malformations, (ii) intrauterine death (IUD), (iii) cystic hygroma and hydrops fetalis, (iv) isolated abnormalities of amniotic fluid, and (v) intrauterine growth restriction (IUGR). The highest diagnostic yield was in fetuses with isolated abnormalities of amniotic fluid (77%), followed by those with IUGR (75%), with IUD (69.6%), those in group five (55.2%) and lowest (45%) in fetuses with malformations (P < 0.001). A cause was identified in 77.8% fetuses with multiple malformations compared to 37.5% with isolated malformations (P < 0.001), with chromosomal abnormalities in 31.8% versus 9.9% respectively (P < 0.001) and malformation syndromes in 42.5% versus 26.3% (P < 0.001). Placental examination provided the highest yield in IUD, IUGR, and oligohydramnios (43.1%; P < 0.003) whereas chromosomal analysis was most useful in cystic hygroma/NIHF (28.9%; P < 0.001). This information on the diagnostic yield in fetal autopsy related its common indications, can be utilized to counsel families of the utility of autopsy to establish cause and recurrence risks and thereby assist then to make an informed decision to consent for the procedure. © 2016 Wiley Periodicals, Inc.

Citrin deficiency: A treatable cause of acute psychosis in adults.

Citrin deficiency is an autosomal recessive genetic disorder caused by a defect in the mitochondrial aspartate/glutamate antiporter, citrin. The disorder manifests either as neonatal intra-hepatic cholestasis or occurs in adulthood with recurrent hyperammonemia and neuropsychiatric disturbances. It has a high prevalence in the East Asian population, but is actually pan-ethnic. We report the case of a 26-year-old male patient presenting with episodes of abnormal neuro-psychiatric behavior associated with hyperammonemia, who was diagnosed to be having citrin deficiency. Sequencing of the SLC25A13 gene revealed two novel mutations, a single base pair deletion, c. 650delT (p.Phe217SerfsFNx0133) in exon 7, and a missense mutation, c. 869T>C (p.Ile290Thr) in exon 9. Confirmation of the diagnosis allowed establishment of the appropriate management. The latter is an essential pre-requisite for obtaining a good prognosis as well as for family counseling.

Genetic Testing in Pediatric Epilepsy: Tools, Tips, and Navigating the Traps.

With the advent of high-throughput sequencing and computational methods, genetic testing has become an integral part of contemporary clinical practice, particularly in epilepsy. The toolbox for genetic testing has evolved from conventional chromosomal microarray and epilepsy gene panels to state-of-the-art sequencing techniques in the modern genomic era. Beyond its potential for therapeutic benefits through precision medicine, optimizing the choice of antiseizure medications, or exploring nonpharmacological therapeutic modalities, genetic testing carries substantial diagnostic, prognostic, and personal implications. Developmental and epileptic encephalopathies, the coexistence of neurodevelopmental comorbidities, early age of epilepsy onset, unexplained drug-refractory epilepsy, and positive family history have demonstrated the highest likelihood of yielding positive genetic test results. Given the diagnostic efficacy across different testing modalities, reducing costs of next-generation sequencing tests, and genetic diversity of epilepsies, exome sequencing or genome sequencing, where feasible and available, have been recommended as the first-tier test. Comprehensive clinical phenotyping at the outset, corroborative evidence from radiology and electrophysiology-based investigations, reverse phenotyping, and periodic reanalysis are some of the valuable strategies when faced with inconclusive test results. In this narrative review, the authors aim to simplify the approach to genetic testing in epilepsy by guiding on the selection of appropriate testing tools in the indicated clinical scenarios, addressing crucial aspects during pre- and post-test counseling sessions, adeptly navigating the traps posed by uncertain or negative genetic variants, and paving the way forward to the emerging testing modalities beyond DNA sequencing.

Role of next generation sequencing in diagnosis and management of critically ill children with suspected monogenic disorder.

Next generation sequencing based diagnosis has emerged as a promising tool for evaluating critically ill neonates and children. However, there is limited data on its utility in developing countries. We assessed its diagnostic rate and clinical impact on management of pediatric patients with a suspected genetic disorder requiring critical care. The study was conducted at a single tertiary hospital in Northern India. We analyzed 70 children with an illness requiring intensive care and obtained a precise molecular diagnosis in 32 of 70 probands (45.3%) using diverse sequencing techniques such as clinical exome, whole exome, and whole genome. A significant change in clinical outcome was observed in 13 of 32 (40.6%) diagnosed probands with a change in medication in 11 subjects and redirection to palliative care in two subjects. Additional benefits included specific dietary management (three cases), avoidance of a major procedure (one case) and better reproductive counseling. Dramatic therapeutic responses were observed in three cases with SCN1A, SCN2A and KCNQ2-related epileptic encephalopathy. A delayed turn-around for sequencing results was perceived as a major limiting factor in the study, as rapid and ultra-rapid sequencing was not available. Achieving a precise molecular diagnosis has great utility in managing critically ill patients with suspected genetic disorders in developing countries.

A Rare Case of Mosaic 3pter and 5pter Deletion-Duplication with Autism Spectrum Disorder and Dyskinesia.

Introduction: There is evidence that neurodevelopmental disorders are associated with chromosomal abnormalities. Current genetic testing can clinch an exact diagnosis in 20-25% of such cases. Case Description. A 3 years and 11 months old boy with global developmental delay had repetitive behaviors and hyperkinetic movements. He was stunted and underweight. He had ataxia, limb dyskinesia, triangular face, microcephaly, upward slanting palpebral fissure, hypertelorism, retrognathia, posteriorly rotated ears, long philtrum, thin lips, broad nasal tip, polydactyly, tappering fingers, and decreased tone in the upper and lower limbs with normal deep tendon reflexes. Magnetic resonance imaging of the brain, ultrasound of the abdomen, and ophthalmological evaluation were normal. Brain evoked response auditory revealed bilateral moderate hearing loss. He fulfilled the Diagnostic Statistical Manual 5 criteria for autism. In the Vineland Social Maturity Scale, his score indicated a severe delay in social functioning. His genetic evaluation included karyotyping, fluorescence in situ hybridization (FISH), and chromosomal microarray analysis (CMA). The karyotype report from high-resolution lymphocyte cultures was mos 46, XY, der(3)t(3; 5)(p26; p15.3)[50]/46, XY,der(5) t(3;5) (p26;p15.3)[50].ish. His karyotype report showed a very rare and abnormal mosaic pattern with two cell lines (50% each). Cell-line#1: 3pter deletion with 5pter duplication (3pter-/5pter+) and cell-line#2: 3pter duplication with 5pter deletion (3pter+/5pter-) derived from a de novo reciprocal translocation t(3; 5)(p26; p15.3) which was confirmed by FISH. The chromosomal microarray analysis report was normal. The two cell lines (50% each) seem to have balanced out at the whole genome level. Occupational, sensory integration, and behavior modification therapy were initiated for his autistic features, and anticholinergic trihexiphenidyl was prescribed for hyperkinetic movements. Conclusion: This case highlights a rare genetic finding and the need for timely genetic testing in a child with dysmorphism and autism with movement disorder to enable appropriate management and genetic counselling.

The molecular landscape of oculocutaneous albinism in India and its therapeutic implications.

Oculocutaneous albinism is an inherited disorder of melanin biosynthesis, characterized by absent or reduced pigmentation of the skin, hair, and eyes. Molecular alterations of genes that cause non-syndromic albinism in Asian Indians are poorly characterized. This information would be useful for developing therapies for this disorder. We analyzed 164 persons with non-syndromic albinism, belonging to unrelated families from all parts of India, for molecular changes in the causative genes. Subjects with white hair, white skin, and red iris had their tyrosinase gene sequenced and were also tested by MLPA for deletions/duplications. Subjects with negative results or with darker skin, golden/brown or darker hair had sequencing of TYR, P, TYRP1, SLC45A2 and GPR143 genes. Pathogenic variants in TYR (OCA1) were observed in 139 (84.7%) patients, in the P gene (OCA2) in 20 (12.2%), in TYRP1 (OCA3) in two (1.2%), in SLC45A2 (OCA 4) in one (0.61%), and in GPR143 (X-linked ocular albinism) in two (1.2%) patients. Of 278 alleles with variants in TYR, 179 (64.3%) alleles had (p.R278*) alteration, suggesting the possibility of therapy with a stop codon readthrough molecule. We report 20 patients with 13 disease associated variants in the P gene and 18 novel pathogenic variants in TYR, P, TYRP1, SLC45A2 and GPR143 genes. The data are compared with those reported from India, Pakistan and rest of the world. The therapeutic options in albinism are briefly described, opening this field for future therapies.

Osteopathia Striata with Cranial Sclerosis: A Face-to-Radiograph-to-Gene Diagnosis.

Osteopathia striata with cranial sclerosis is an X-linked dominant bone dysplasia with osteosclerosis. It should be suspected in girls with macrocephaly, intellectual disability with unique facial dysmorphic features. We described the clinical and radiological profile of a patient with this rare disorder. A novel heterozygous variant was identified in the AMER1 gene which leads to premature truncation of the AMER1 protein. Facial gestalt recognition using artificial intelligence and radiographic features were used to narrow the differential diagnosis.

Clinical and Genetic Profile of Children With Short Stature Presenting to a Genetic Clinic in Northern India.

OBJECTIVE: To define the spectrum of genetic disorders in patients with short stature visiting the genetic out-patient department in a tertiary care hospital. METHODS: A chart review was done for 455 individuals (10 months-16 yrs) with short stature, who were evaluated at the genetic clinic from 1 January, 2017 upto 31 October, 2018. 226 patients who needed detailed evaluation, the spectrum of genetic diagnosis is presented. RESULTS: Proportionate short stature was identified in 63% individuals (n=142) of which 93 (65%) were recognizable syndromes such as Turner syndrome, and William syndrome, and RASopathies. In clinically undefined syndromes (39, 27%), a diagnosis could be made by karyotype (n=3/10), chromosomal microarray (6/12) and exome sequencing (1/6). In the 84 children in the disproportionate short stature group (37%), lysosomal storage disorders (LSDs) (45%, n=38) were identified by enzyme analysis in 86.8% and skeletal dysplasias (44%, n=37) identified by skeletal survey in 89% cases. CONCLUSIONS: In undefined syndromic short stature, chromosomal microarray may be the first investigation of choice if phenotyping is not suggestive of a specific genetic syndrome. Exome sequencing can be useful in identifying newer genes among idiopathic and familial short stature cohorts.

Hypervitaminosis D and Acute Interstitial Nephritis: Tale of Injections.

A 33-year-old man came with nausea, vomiting and abdominal pain due to hypercalcaemia and renal dysfunction following two doses of intramuscular vitamin D injections. Levels of vitamin D were repeatedly above 300 ng/ml over a period of 10 months. Whole-body PET CT scan revealed a thin-walled collection in the right gluteal region. The patient refused a surgical intervention for the same. After 7 months of follow-up, the abscess ruptured spontaneously and was then surgically debrided. At this point, a history of pentazocine addiction was uncovered. One month later, vitamin D levels began to fall along with improvement in serum calcium and creatinine. This case unravels a diagnostic odyssey which ended with a simple surgical debridement. We aim to highlight that vitamin D supplementation in 'megadoses' in the presence of active infection can have an exaggerated response and may take months to resolve.